How Change In Secondary Structure Affects Proteins

• Periodical Listing
• IUCrJ
• v.two(Pt half-dozen); 2015 November 1
• PMC4645109

IUCrJ. 2015 Nov 1; 2(Pt half-dozen): 643–652.

Changes in protein structure at the interface accompanying complex formation

Devlina Chakravarty

^aSection of Biochemistry, Bose Institute, P-1/12 CIT Scheme VIIM, Kolkata 700 054, Republic of india

Joël Janin

^bIBBMC, CNRS UMR 8619, Universite Paris-Sud 11, Orsay, France

Charles H. Robert

^cCNRS Laboratoire de Biochimie Theorique, Institut de Biologie Physico-Chimique (IBPC), Universite Paris Diderot, Sorbonne Paris Cité, 13 Rue Pierre et Marie Curie, 75005 Paris, French republic

Pinak Chakrabarti

^aDepartment of Biochemistry, Bose Plant, P-i/12 CIT Scheme VIIM, Kolkata 700 054, Republic of india

Received 2015 Jun 12; Accepted 2015 Aug 16.

Abstruse

Protein interactions are essential in all biological processes. The changes brought about in the structure when a gratuitous component forms a circuitous with another molecule need to exist characterized for a proper agreement of molecular recognition besides as for the successful implementation of docking algorithms. Hither, unbound (U) and bound (B) forms of protein structures from the Protein–Protein Interaction Affinity Database are compared in society to enumerate the changes that occur at the interface atoms/residues in terms of the solvent-attainable surface surface area (ASA), secondary structure, temperature factors (B factors) and disorder-to-order transitions. It is constitute that the interface atoms optimize contacts with the atoms in the partner poly peptide, which leads to an increase in their ASA in the bound interface in the bulk (69%) of the proteins when compared with the unbound interface, and this is independent of the root-mean-square deviation betwixt the U and B forms. Changes in secondary structure during the transition indicate a likely extension of helices and strands at the expense of turns and coils. A reduction in flexibility during complex formation is reflected in the decrease in B factors of the interface residues on going from the U grade to the B class. At that place is, however, no distinction in flexibility between the interface and the surface in the monomeric construction, thereby highlighting the potential problem of using B factors for the prediction of binding sites in the unbound class for docking another protein. sixteen% of the proteins have missing (disordered) residues in the U form which are observed (ordered) in the B form, mostly with an irregular conformation; the information gear up also shows differences in the composition of interface and non-interface residues in the disordered polypeptide segments as well as differences in their surface burying.

Keywords: protein–protein interactions, protein flexibility, disorder–order transition, bound and unbound protein forms, interface area, crystallographic temperature factor, secondary structure, bioinformatics, molecular recognition

one. Introduction

Poly peptide–protein recognition plays a crucial part in many biological processes, including Dna replication, protein degradation, signal transduction and metabolic processes (Stites, 1997 ▸). Interaction with small molecules, nucleic acids and other proteins takes place through bounden at specific sites. Protein–protein interactions have distinct characteristics, such equally directly physical contact, surface complementarity and a specific, well defined interface. Protein structure, dynamics and function are interdependent. Relating construction to dynamics and office is essential in agreement molecular-recognition processes (Boehr et al., 2009 ▸; Mittag et al., 2010 ▸; Tompa & Fuxreiter, 2008 ▸). Factors such every bit hydrophobicity and the specific location of residues capable of forming hydrogen bonds and electrostatic interactions help to distinguish the interface from the rest of the surface (Janin et al., 2008 ▸). Jones and Thornton, among others, have discussed the features of interface patches, for case residue propensity, planarity, surface accessibility and protrusion, that make them different from the rest of the protein surface (Jones & Thornton, 1995 ▸, 1997 ▸). Conservation of interface residues and their clustering can also be used as a discriminating factor (Guharoy & Chakrabarti, 2010 ▸). Interface residues undergo more significant conformational changes than other surface residues and have evolved to retain the specificity of their interactions (Rajamani et al., 2004 ▸). Although many proteins interact as quasi-rigid bodies, undergoing very niggling conformational alter every bit they form complexes, in many cases the conformational change is pregnant and may or may not be restricted to the interface region (Swapna et al., 2012 ▸). Proper integration of these changes is important for estimating the bounden affinity between ii proteins from structural data (Janin, 2014 ▸), still incorporating them is the primary bottleneck in the development of robust protein–protein docking algorithms (Aloy et al., 2005 ▸; Bonvin, 2006 ▸). There have already been attempts to decipher the changes associated with the transition of the gratis (or unbound) form of a molecule to the bound course in the circuitous, for example involving side-concatenation conformations (Guharoy et al., 2010 ▸; Ruvinsky et al., 2011 ▸).

Analyses of protein–poly peptide interfaces are usually performed on complexes that are bachelor every bit crystal structures. Still, proteins are dynamic and be in ensembles of interchanging structures. Binding to small molecules, nucleic acids or other proteins leads to shifts in the populations of these conformers; the term conformational change is a autograph for such functional shifts. The affinity in item depends on both the unbound-country and bound-state ensembles. To improve understand the changes brought virtually by association, we accept used the protein–protein binding-affinity benchmark, which is a nonredundant set up of 144 complexes for which loftier-resolution structures are available for both the complexes and their unbound components and for which dissociation constants take been measured using biophysical techniques (Kastritis et al., 2011 ▸). We have recently looked at the changes in attainable surface area of interface atoms in pairs of unbound (U) and bound (B) forms of proteins (Chakravarty et al., 2013 ▸); in this piece of work, we extend these analyses and also written report the changes in secondary structures, temperature factors (B factors) and disorder-to-guild transitions.

2. Materials and methods

We used the Poly peptide–Poly peptide Interaction Affinity Database (Kastritis et al., 2011 ▸) containing 144 complexes along with the respective unbound structures, except for seven antibiotic–antigen complexes for which the unbound structure is not known. We thus considered 281 spring–unbound construction pairs. The unbound structure is designated U and the bound structure (isolated from its partner component) is designated B. The bound form of the component in the presence of its partner (i.e. in circuitous) is designated C. EMBOSS (Rice et al., 2000 ▸) was used to perform the local alignment (using the Smith–Waterman algorithm) of the polypeptide bondage constituting U/B pairs; 249 had a sequence identity of ≥96%, with the rest having values in the range 90–95%. Based on the sequence alignment, the interface residues as seen in the circuitous were mapped onto those in the unbound state using ProFit (McLachlan, 1982 ▸) and Biopython (Cock et al., 2009 ▸). Amino acids differing between the jump and unbound protein sequences (Supplementary Table S1a), and positions at which data are missing attributable to order–disorder transitions in the 2 PDB files nether consideration (Supplementary Tabular array S1c), were excluded from structural analysis. At that place are 27 structures with different balance names in U and B affecting forty residues. Any modified residues (Supplementary Tabular array S1b) were manually edited to match the natural amino acid. Interface atoms were identified as those losing more 0.one Åⁱⁱ of surface surface area upon circuitous formation (B to C; Chakrabarti & Janin, 2002 ▸). NACCESS (Hubbard, 1992 ▸) was used for the adding of solvent-accessible surface area (ASA). Equally discussed in Chakravarty et al. (2013 ▸), one has to consider the ambiguity in cantlet labels (specially of aromatic residues) while calculating the surface area cached in going from U to B. Hydrogen bonds were assigned using HBPLUS (McDonald & Thornton, 1994 ▸) with default geometrical parameters. Secondary structures were determined using DSSP (Kabsch & Sander, 1983 ▸).

In the affinity information set up, 17 unbound structures were determined past NMR, of which 13 accept multiple models or conformations. For each of them the beginning model was used. However, in a control calculation, the surface parameters calculated using all of the models were found to be essentially the same as those employed in the reported calculations.

2.ane. Terms and equations used to describe the changes in the interface

• (i) ΔASA = [ASA(B) – ASA(U)], where ASA(B) is the solvent-accessible surface area of the mapped interface atoms in the bound land and ASA(U) is the value in the unbound state. In this calculation merely the interface atoms nowadays in both states were used. This is distinct from the cached surface area, BSA = [ASA(B) – ASA(C)], calculated in the standard way using all of the interface atoms.

• (ii) δA = ΔASA/ASA(B) is the divergence in ASA relative to the total value in the bound state.

• (iii) bf _r ′ = [bf _r − μ(bf)]/σ(bf), where bf _r is the boilerplate B factor of the C, C^α, O, N and C^β atoms of balance r (C, C^α, O and North for Gly) and μ(bf) and σ(bf) are the mean and standard deviation of the B factors for that concatenation, respectively. The normalized bf _r ′ values were used to derive the averages over the interface, surface and core and rim regions of the interface (Chakrabarti & Janin, 2002 ▸).

• (iv) D = ; the Euclidean altitude D was used to quantify the modify in n_south , the percentage limerick of the secondary-structure type south of interface residues, between the spring (northward ^B _s ) and the unbound (n ^U _s ) forms; the m = 4 secondary-structure types are defined as helix, strand, turn and whorl.

• (v) The Euclidean metrics, Δb, for the B factors of residues in different states/structural regions were calculated in a like fashion, Δb = , where n represents the number of amino-acid types and bf _i ^(ane) and bf _i ⁽²⁾ are the scaled B factors of residue type i in states one and 2, respectively. United states that were compared were interface, non-interface, jump and unbound.

3. Results

3.1. Modify in the ASA of interface atoms on going from the U country to the B state

Previously, we had shown that on going from the U form to the B form the interface atoms undergo an increase in accessible surface area (ASA), leading to a positive δA value (Supplementary Fig. S1; mean = three.3 ± 9.two%), which is the result of conformational changes taking place at the interface (Chakravarty et al., 2013 ▸). (As a control, nosotros checked the variation of the ASA of free surface residues, which show only an insignificant increase, with a mean value of 0.ninety ± 6.06%.) Considering the whole rest, which includes not-interface atoms, the increase can still be seen (1.three ± 8.03%) merely is smaller than that exhibited by the interface atoms solitary. The ASA increase reflects what might be called a 'partner attraction result': interface atoms are extended in the bound state to optimize contact with the binding partner. In addition to maximizing van der Waals interactions, the increment in the ASA of interface atoms could also be the effect of optimizing interchain hydrogen-bond geometry. As a simple quantification of this, we used structures for which the combined r.chiliad.s.d. for the U-to-B change for the two components (I_r.m.southward.d. according to Kastritis et al., 2011 ▸) is <one Å. For these 59 cases we generated the pseudo-complex by superimposing the two U forms onto the corresponding B structures. The average number of hydrogen bonds in the pseudo-circuitous is 3.seven ± 2.five, whereas in the real complex information technology is 8.0 ± 3.7, a 45% increase. An example of the local adjustment of the two U structures leading to the germination of a hydrogen bond in the circuitous is shown in Fig. 1 ▸: the structural rearrangement pulls out the Tyr rest such that there is a net gain in ΔASA.

Hydrogen-bail geometries (distances shown) in α-amylase (green) and tendamistat (cyan) between His201 NE2 and Tyr820 OH for (a) the pseudo-complex and (b) the experimental complex [PDB entry 1bvn (Wiegand et al., 1995 ▸); PDB entries 1pig (Machius et al., 1996 ▸) and 1hoe (Pflugrath et al., 1986 ▸) are the U forms]. ΔASA for the participating atom and all of the interface atoms of the residues are −0.six and −3.2 Ų, respectively, for His, and 4.two and 15.5 Ų, respectively, for Tyr.

While the majority of complexes show an increase in ASA, 31% (88 of the 281 components) have a negative δA value, indicating that the interface atoms are pulled dorsum into the structure to facilitate the interaction with the incoming partner molecule: a 'partner accommodation' effect. Fig. 2 ▸ shows such an example with a δA value (−10%) from the opposite side of the distribution. It is seen that for the core domain of the HspBP1 poly peptide the effect of binding has been to pull the interface atoms, which were extended into the solvent in the U class, towards itself to allow a closer approach past the partner molecule (the Hsp70 ATPase domain). While the two component contributions in the complex are weakly correlated (Fig. 3 ▸), we note that the proportions of complexes in which the ΔASA contributions of the two partner proteins are both positive (65 complexes; 47%), both negative (15 cases; 11%) and mixed positive and negative (57 complexes; 42%) are consistent with a simple statistical model of independent component contributions (p ² ₊ = 47%, p ² ₋ = ten% and iip ₊ p ₋ = 43% for p ₊ = 0.69 and p ₋ = 0.31). Thus, the 'partner accommodation upshot' does not usually operate simultaneously on both components, and circuitous formation is usually accompanied by the 'partner allure effect'. ΔASA has a poor correlation with interface r.chiliad.s.d. and BSA (Supplementary Fig. S2), indicating it to exist substantially independent of the size of the interface or the root-mean-foursquare departure of the interface atoms.

The complex between the core domain of HspBP1 and the Hsp70 ATPase domain, an example of the modify in the position of interface residues (stick representation; scarlet in the B form and bluish in the U form). Protein bondage are shown in drawing representation in greenish for the B course (PDB entry 1xqs) and in pink for the U form (PDB entry 1xqr) of the core domain of HspBP1 (Shomura et al., 2005 ▸) containing the labelled interface residues; the other component (the Hsp70 ATPase domain) in the B form is shown in cyan. ΔASA = −175 Ų and δA = −10%. The ΔASA values for the interface atoms of the residues shown are −43 Ų for Arg217, −twenty Å^two for Glu218 and −16 Å^two for Phe210.

Plot of ΔASA of the interface atoms separated into the two components for each complex. The greater of the two values is labelled ΔASA2 and the lesser ΔASA1.

three.2. Changes in secondary structure

The change in the per centum composition of secondary-structural elements for the U to B transition was calculated, and 76% cases (213 of 281) showed some change. To restrict the analysis to meaningful changes, we computed the Euclidean distance (D) between the compositions of the four structural elements. The boilerplate value of D is five.half dozen (±v.4), and we used structural pairs with D > 5 (134 cases) to sympathize the structural changes accompanying complex germination (Fig. iv ▸ a) [the histograms for D > ten and D > 15 (Supplementary Figs. S3a and S3b) look very similar]. It tin can be seen that complex formation leads to an increase in helical and strand content (especially the sometime) at the expense of irregular (and to some extent plough) regions in the structure. 91 structural pairs show an irregular/turn (C/T) to helix/strand (H/S) transition, affecting 75 helices and 81 strands, corresponding to 34% of helices and 38% of strands, respectively, of these structural elements in the B form of the proteins. These cases take an average D value of 7.8 ± 4.9, with 224 residues irresolute conformation. The majority of these (161 cases) are involved in the extension of an already existing helix or strand (Fig. 4 ▸ b). Cases of extension seem to marginally favour the C-last end of helices and the N-terminal stop of strands (Supplementary Fig. S3c). The residues located in the interface core (108 of 224; 48%) and rim (52%) are affected equally, among which Arg, Glu, Ser and Tyr are those more frequently involved in the transition from C/T to H/S. 2 representative examples showing secondary-structural changes are presented in Fig. 5 ▸.

Secondary-structural changes during the U-to-B transition. (a) The modify in percent composition between the ii states (B – U) for the secondary-structural elements (helix, H; strand, S; plough, T; irregular, C) for the cases with Euclidean distances between the two sets of compositions of >5. (b) Percentage composition of 224 residues showing the C/T to H/South transition, categorized into the extension of an already existing helix/strand (EH and ES) or the formation of a new helix/strand (FH and FS).

Examples showing changes in secondary-structural elements (left console, U; right panel, B). Stretches in the interface are in yellowish. (a) Amicyanin (PDB entry 2rac; Zhu et al., 1998 ▸) in complex (PDB entry 2mta; Chen et al., 1994 ▸) with methylamine dehydrogenase exhibits the formation of two antiparallel β-strands (Pro52, Asn54, His56 and Val58 in 1, and Lys68, Gly69, Pro70, Met71 and Lys73 in the other) and North-terminal (Arg99) and C-last (His91) extension of two other strands. (b) Metalloproteinase inhibitor 1 (PDB entry 1d2b; Wu et al., 2000 ▸) in complex (PDB entry 2j0t; Iyer et al., 2006 ▸) with MMP1 intersitial collagenase displays the formation of a small helix (Glu67, Ser68, Val69 and Cys70) and C-concluding (Lys88) extension of a strand.

3.iii. Assay of missing residues in the unbound form

95 proteins of the 281 have one or more than interface atoms missing in the crystal structure of the U state that are present in the B state. 46 proteins (16%) accept interface residues missing in the unbound form (on boilerplate iv missing residues per component). We will refer to these atoms and residues as 'missing' even though they are clearly present in the bound form. Missing atoms constituted ∼iv% of the total interface atoms in the data set and 12.5% for the 95 structures, with the most extreme beingness MAPKAP kinase 2 (PDB entry 3fyk; Anderson et al., 2009 ▸), in which 52% of the interface atoms (39 of 80 interface residues) are missing. Usually the interface and non-interface residues are interspersed in a given stretch of missing residues, and in that location are 34 such cases (12%) with two or more missing residues, the sequences of which can exist seen in Supplementary Table S2. Such regions undergo a disorder-to-gild transition upon bounden to their interacting partner, and have special importance in elucidation of the structure–function relationships of proteins (van der Lee et al., 2014 ▸). Interestingly, in more than one-half of the cases the missing segment is at the polypeptide concatenation termini.

The statistics for the missing residues in the U form are provided in Table i ▸. For all of the missing stretches of amino acids given in Supplementary Table S2, ii values are reported: ane concerning simply the interface residues and the other only the non-interface residues. The composition of missing residues is like to that of the interface equally a whole, although aromatic residues (Phe, Tyr and Trp) seem to have a lower tendency to exist disordered. Relative to the total number of missing interface residues, amid the charged residues the longer ones are found in greater numbers (Glu > Asp, Arg > Lys or His). Interestingly, the composition of nonpolar interface residues (Ala, Leu and Ile) is too on the higher side. Thus, the hydrophobic effect would appear to have a role in determining which residues in the matted regions contribute to binding. It may be noted that intrinsically disordered proteins are known to expose their few hydrophobic residues for interaction with the partner (Mészáros et al., 2007 ▸).

Table ane

Statistics for interface residues missing in the U grade and their secondary structure in the B form

		% relative to full No. of		Secondary structure in the B class of residues missing in U† (%)
Balance	No. missing‡	Interface residues of the same type	Missing residues‡	H	South	T	C
Ala	15 (v)	4.7	7.half-dozen (half-dozen.1)	27 (35)	0	xiii (x)	60 (55)
Arg	14 (2)	3.2	vii.1 (4.ix)	28.half dozen (25)	0	21.4 (eighteen.8)	50 (56.ii)
Asn	thirteen (4)	iii.0	6.6 (5.two)	7.7 (five.9)	0	46.2 (41.1)	46.ii (52.9)
Asp	x (10)	2.3	v.5 (half dozen.i)	50 (xxx)	0	0 (25)	l (45)
Cys	1 (2)	0.6	0.5 (0.9)	0	0	0 (33.3)	100 (66.7)
Gln	nine (5)	ii.five	4.5 (4.3)	11.1 (vii.1)	0	33.iii (fifty)	55.6 (42.9)
Glu	18 (10)	iii.8	9.one (eight.half-dozen)	11.1 (14.3)	five.6 (3.six)	27.8 (32.1)	55.six (50)
Gly	13 (12)	2.5	6.6 (vii.7)	0	vii.vii (4)	23.1 (forty)	69.2 (56)
His	7 (3)	3.2	3.5 (iii.i)	0 (10)	0	57.1 (l)	42.9 (40)
Ile	10 (11)	three.5	six.i (6.4)	10 (9.5)	0 (4.8)	10 (19)	80 (66.seven)
Leu	15 (four)	3.3	7.6 (5.8)	33.three (31.half-dozen)	0	six.7 (five.three)	60 (63.2)
Lys	11 (7)	2.iii	five.6 (5.v)	27.4 (27.8)	0 (5.6)	36.4 (22.ii)	36.4 (44.5)
Met	7 (2)	5.0	3.five (2.8)	57.2 (66.7)	14.3 (xi.1)	28.vi (22.2)	0
Phe	5 (2)	i.9	2.v (ii.2)	0	0	0 (fourteen.3)	100 (85.7)
Pro	11 (9)	iii.vi	5.6 (6.1)	18.2 (ten)	0	9.1 (15)	72.7 (75)
Ser	11 (21)	two.1	5.half-dozen (9.viii)	27.3 (12.5)	0 (six.25)	36.four (37.five)	36.4 (43.8)
Thr	seven (13)	1.5	iii.v (vi.one)	28.six (25)	0 (10)	28.6 (10)	42.9 (55)
Trp	iv (ii)	2.half dozen	2.0 (ane.eight)	0 (sixteen.7)	25 (16.7)	0	75 (66.seven)
Tyr	7 (3)	1.6	3.0 (2.5)	0 (10)	0	57.i (forty)	42.9 (l)
Val	9 (4)	2.seven	4.5 (iv.0)	0	xi.1 (15.4)	11.one (vii.7)	77.8 (76.9)
Full	197 (131)		100

We also analyzed the secondary structures adopted in the jump form by these missing residues. In full general, >50% of the missing interface residues adopt an irregular conformation in the jump state; next in level of occurrence are helices and turns, with strands seeming to be the least favoured. Because interface residues located in the terminal peptide segments simply, ane observes a slightly college tendency to adopt an irregular conformation (65%, as opposed to 20% in T and 15% in H). Met and Leu assume a helical conformation in greater percentages, as tin be expected from the mostly higher preference of these residues for this type of secondary structure.

Of the missing residues, 49% become core interface residues (those which have atoms fully buried in the interface); withal, these contribute 70% of the BSA; the BSA values of core residues missing in U exceed those of the rim with a P value of 0.01. These observations are in conformity with a previous report (Chakrabarti & Janin, 2002 ▸). In the 46 proteins missing one or more interface residues, the latter contribute 17% of the BSA in the bound state (190 ± 278 Ų of 1017 ± 568 Ų); however, the distribution is rather big, ranging from ∼57% in the MAPK-activated protein kinase 2 (MK2) part of the assembly formed with p38 (PDB entries 3fyk and 2oza; Anderson et al., 2009 ▸; White et al., 2007 ▸) to 0.i% in the complex formed past Ran-specific GTPase-activating protein with GTP-binding nuclear protein RAN (PDB entries 1yrg and 1k5d; Hillig et al., 1999 ▸; Seewald et al., 2002 ▸), with 39 and one residues, respectively, missing in the U land.

In the 95 structures with missing atoms, each missing atom contributes 11.5 ± six.eight Å^two to the BSA in the bound state, while the remaining ('non-missing') atoms each contribute nine.four ± one.five Ų (P = 3 × x⁻²¹ for the two populations with 977 and 6945 atoms, respectively). Thus, at the local level the interface elements that undergo a disorder-to-social club transition upon forming the circuitous bury somewhat more surface than other atoms in the interface. However, at the database level the issue is more marked; structures for which no interface residues are missing in the U form bury on average 786 ± 336 Ų in the circuitous, while structures with i or more interface residues missing bury more: 1017 Ų, every bit given above (P = 0.005). Indeed, when calculated for the structures missing five residues or more (13 structures in all), surface burial is larger still: 1507 ± 795 Å^two (P = 0.003). The presence of missing residues is thus seen to be associated with a larger degree of surface burial upon forming the complex, similar to systems undergoing a conformational change upon association (Kastritis et al., 2011 ▸); both effects presumably bespeak compensation of the corresponding free-energy punishment.

The total interface in a complex is made up of contributions from both components, the BSA values of which are generally similar but not equal. It is of involvement to study the contribution of a specific balance not but to the BSA of its own component ('parent'), merely too to that of the partner component owing to their interaction. Overall, the missing residues in a given component contribute 155 ± 270 Ų to the BSA of the parent and virtually the same (156 ± 246 Ų) to that of the partner. Nevertheless, in those cases for which missing residues contributed more than than 200 Å^two to the parent (13 structures, missing nine residues per structure on average), the contribution to the partner was smaller on boilerplate by 34 ± 50 Å^two. It may be mentioned in connexion that the BSA values of the interacting proteins are unremarkably nearly identical; in the instance of protease–inhibitor complexes, nonetheless, the convex nature of the inhibitor surface plumbing equipment into the concave active site results in its BSA exceeding that of the enzyme in the ratio 54:46 (Lo Conte et al., 1999 ▸). Figs. 6 ▸ and 7 ▸ provide two illustrations of a missing segment and the structure (more often than not of irregular conformation) adopted in the B grade. The example in Fig. 6 ▸ is a instance in which the gain in BSA from missing residues in the parent molecule exceeds that in the partner by 139 Åⁱⁱ. The asymmetric nature of the BSA values for the two sides is owing to the improve fitting of the disordered residues into the grooves and crevices of the more ordered interacting partner. Favourable interactions arising from the burying of these residues should likewise help to compensate for the entropic loss of ordering them.

The construction of the interface formed in homo tissue inhibitor of metalloproteinases 2 when it forms a circuitous with type IV collagenase (PDB entry 1gxd; Morgunova et al., 2002 ▸); the inhibitor is denoted in cyan and the enzyme in violet. Surface representations of the proteins are displayed. The U state (PDB entry 1br9; Tuuttila et al., 1998 ▸) is non shown hither. The interface residues are split into ii categories: the residues missing in the unbound structure are in blue and those seen in both the U and B forms are in orangish. The missing segment (183–192) is composed of both non-interface residues (shown in red) and interface residues (blueish). The missing residues contribute 504 Åⁱⁱ to the BSA of 1268 Ų of the inhibitor.

The loop (i–12) missing in the U form of neurotrophin-4 (PDB entry 1b98; Robinson et al., 1999 ▸; shown every bit greenish cartoon) which is present in the B grade (PDB entry 1hcf; Banfield et al., 2001 ▸; cyan) on forming a circuitous with the BDNF/NT-3 growth factor receptor TrkB-d5 (magenta drawing). The interface residues (in bluish) are interspersed with not-interface residues (in red) in the missing loop. The contribution of the missing residues is 383 Ų to the BSA of 765 Åⁱⁱ.

The rest composition does not modify much if we consider the not-interface residues in the missing stretches of amino acids (Table ane ▸); forth with the charged residues (Glu, Asp, Arg, Lys), Ser, Thr and Gly are seen to occur in large numbers also. This is in accord with what is observed in intrinsically disordered proteins, which are enriched in charged and polar amino acids and depleted in beefy hydrophobic groups (van der Lee et al., 2014 ▸). The missing stretches exhibit similar features as the interface residues within them, usually taking up turn or irregular conformations in the bound structure.

3.4. Comparison of B factors

The crystallographic B factor (temperature factor or atomic readapt­ment parameter) is a measure of the oscillation of an atom around its mean position owing to thermal motion and positional disorder. Normalized B factors have been used to compare structures (Parthasarathy & Murthy, 2000 ▸). Information technology has been recognized that residues in the interface have lower B factors than those in the protein exterior (Jones & Thornton, 1995 ▸), suggesting that residues participating in protein–protein interactions are less flexible than those on the gratuitous surface. This inference was based on an analysis of complex structures merely. All the same, a subsequent comparison of 57 monomeric structures with their bound homologues (>70% sequence identity) indicated that even in the unbound state the distribution of B factors is somewhat lower for the interface than for the rest of the surface (Neuvirth et al., 2004 ▸). Along these lines, we take compared the interface and the surface residues for the bound every bit well as the unbound structures.

The scaled hateful B factor of the courage atoms C, C^α, O and Northward (along with C^β for non-Gly residues) were calculated along with the boilerplate values for each residue type in the interface and the surface regions for both the U and B states. As expected, the boilerplate B factor was observed to be greater for the surface compared with the interface in the B structures (P value < two × x⁻¹⁶; Supplementary Table S3, Fig. 8 ▸ a). Indeed, the normalized B factors for all of the residues in the interface are negative (below the average value for all of the residues in the structure). In dissimilarity, in the unbound structures the interface residues mostly have positive values and, as expected, the values are higher than those observed in the bound interface. Thus, on going from the the U state to the B state the interface residues showroom a desperate reduction in B factor. Although the changes are not as strong, overall the opposite trend was observed for the surface residues (P value = 0.04). Once more applying a Euclidean metric, here divers using the average B factors of amino-acid residues in the two regions of the poly peptide structures and in the ii states (Supplementary Table S3), we find that the maximum changes occur in the interface region as the complex is formed and betwixt the interface and the surface regions in the complex. Overall, the B factors in U are quite similar between the interface and the surface. Interestingly, all the same, hydrophobic residues (notably the aromatic residues) tend to be more flexible at the interface compared with the surface in the U state, while the contrary seems to be the case for polar residues. Grouping Ile, Leu, Met, Phe, Trp and Tyr as nonpolar and Arg, Asn, Cys, Gln, Glu, Gly, His, Lys, Ser and Thr as polar, the difference in B factors is pregnant (the P values are 0.05 and 0.048, respectively). Information technology has been noted that the δA values are higher (>4%) for all of the nonpolar residue types (Chakravarty et al., 2013 ▸). The college flexibility in the U land of the nonpolar residues in the region that would constitute the interface (in B) may thus predispose them to conformational changes accompanying complex formation.

Euclidean distances involving B factors (a) between interface and surface regions (enumerated in Supplementary Table S3) and (b) between interface rim and core regions (Supplementary Tabular array S4) in the U and B states.

The interface residues were further divided into cadre and rim regions (Chakrabarti & Janin, 2002 ▸), and B factors were also compared between these ii regions in the U and B states (Supplementary Table S4). The reduction in B factors is more pronounced in the core region between the two states, which can also be seen from the Euclidean distance between them (Fig. 8 ▸ b); the rim residues show a smaller difference betwixt the 2 forms. This is also reflected in the P values (2 × x^−xvi for the core and 7.602 × ten^−nine for the rim). An analogy of these results for a representative protein is shown in Supplementary Fig. S4. Comparing the core–rim demarcation in Supplementary Fig. S4(e) with the distribution of B factors in the B form (Supplementary Fig. S4b), one can see considerable matching for the core (nighttime blue). At that place is very piddling resemblance to the B factors observed for the U class (Supplementary Fig. S4d). Indeed, there is no significant difference overall between the core and rim residue B factors in the U form (P value = 0.97). This is in contrast to the results from molecular-dynamics simulations, which had indicated a lesser fluctuation of the cadre residues even when the bounden partner is absent (Smith et al., 2005 ▸).

4. Give-and-take

Computation of the accessible surface surface area (ASA) has been very useful in the identification of interface residues (Janin et al., 2008 ▸) and in segregating the interface into cadre, rim and back up regions (Chakrabarti & Janin, 2002 ▸; Levy, 2010 ▸). It has been used to predict the magnitude of binding-induced conformational changes from the structures of either monomeric proteins or bound subunits (Marsh & Teichmann, 2011 ▸). Here, ASA has been used to compare the interface atoms in the unbound and bound states of a protein. Conformational changes brought about past poly peptide–poly peptide interaction are ofttimes discussed in the context of 'induced fit', which however fails to capture the sense of the change observed in the ASA calculations. Ii terms take thus been coined here to distinguish between the increase in the ASA of interface atoms upon complex germination and their decrease: 'partner attraction' and 'partner accommodation' effects, respectively. The former is observed to dominate, although clear examples of the latter are also observed; both are examples of induced fit in the broad sense. In a complex, the interface atoms tend to make fewer contacts within their component as they interact with the other component; information technology is as if the atoms are pulled out of the parent molecule for optimum binding to the partner molecule (Chakravarty et al., 2013 ▸). It has been suggested that the change in side-chain conformation of interface residues may lead to an increase in the relative solvent-accessible surface area on complexation (Ruvinsky et al., 2011 ▸). Nonetheless, rather than at the level of the residue as a whole, we observe that the increase in ASA is more at the level of interface atoms. ΔASA seems to be independent of the overall size of the interface or of whether the molecule binds equally a rigid body or exhibits conformational changes.

Unlike intrinsically disordered proteins (IDPs) made of entirely matted sequences that practice not prefer any tertiary structure in the uncomplexed state, 16% of the proteins considered here contain both structured and disordered regions, which are seen to contribute to poly peptide–protein interaction and thereby perhaps facilitate the regulation of cellular processes (van der Lee et al., 2014 ▸; Dyson & Wright, 2002 ▸; Dunker et al., 2001 ▸; Fong et al., 2009 ▸; Ruvinsky et al., 2011 ▸). Features of matted regions revealed in this work, such equally amino-acid preferences, the amount of ASA buried in complexation etc., may exist general characteristics of IDPs (and intrinsically disordered regions; IDRs), particularly in proteins which function as effectors (van der Lee et al., 2014 ▸; Guharoy et al., 2015 ▸).

Besides the ASA, parameters such every bit hydrogen-bonding patterns, secondary-structure changes and B factors can also be very useful in discerning the interface from the surface. We have seen that in proteins that undergo secondary-structure changes upon the U-to-B transition the most common are extension of the existing helices and strands at the expense of turns and irregular regions (Fig. iv ▸). By defining matted residues equally those with missing coordinates in the crystal structure, i discerns features of disorder-to-order transitions of IDPs or IDRs that play a office in binding. Overall, in going from the U state to the B state proteins adopt a more ordered and regular construction: nonetheless, regions showing a disorder-to-order transition tend to assume more irregular secondary structures in the B land, while parts which were already ordered in the U state tend to shift towards more regular secondary structures, if they modify at all. The ordering of missing residues upon complex formation adds an entropic penalty to the association reaction, which appears to be compensated in part by a greater degree of surface burying in complexes with missing residues, similar to what has been observed for conformational changes (Kastritis et al., 2011 ▸).

Previous studies have demonstrated that in poly peptide–protein complexes (B form) the B factors of the interface residues are lower than those of the surface residues (Jones & Thornton, 1995 ▸; Liu et al., 2010 ▸). In the bound interfaces, we also see that the interface core is significantly less mobile than the rim. In the unbound interfaces, the nonpolar remainder flexibility appears to actually exist somewhat higher than in the non-interface surface, which may be related to these rest types contributing somewhat more to the increase in ASA upon complex formation. Overall, withal, the marked B-factor differences seen in the B grade are of course non present in the U form, in which the interface region is solvent-exposed and mobile. B factors have been used in constructing support vector machines (SVMs) and other popular classifiers to identify protein–protein interaction sites and to distinguish biological interfaces from crystal contacts (Liu et al., 2010 ▸, 2014 ▸; Neuvirth et al., 2004 ▸). Our results signal to limitations in using B factors for the identification of the binding site, especially if one is focusing on the U construction, which is more relevant than the B course in the context of poly peptide–protein complex prediction. Too, forth with other interactions, hydrogen bonding has been used equally a characteristic for predicting protein–protein interaction sites among targets of homologous proteins (Maheshwari & Brylinski, 2015 ▸); nevertheless, the latter is necessarily based on complex structures, which as Fig. 1 ▸ shows cannot exist properly reproduced by the unbound form fifty-fifty for proteins that are considered to comport every bit quasi-rigid bodies when forming the complex.

5. Conclusions

Virtually xc% (122 of 137) of the protein–poly peptide complexes considered in this study show an increase in ASA for interface atoms on going from the U form to the B form for at least one of the two partners, which results from the optimization of contacts through the interface. This change in ASA is contained of whether or not a molecule behaves as a quasi-rigid body or undergoes conformational changes during complex germination. These and other changes that have place in the interface, including optimization of hydrogen bonding to the partner protein, the formation and the extension of regular secondary structures at the price of turns and coils, reduction of B factors (flexibility) and disorder-to-club transitions in the interface residues, presumably contribute to the specificity, the stability and the function of the complex.

6. Related literature

The following references are cited in the Supporting Information for this article: Ratnaparkhi et al. (1998 ▸) and Ševčík et al. (1998 ▸).

Supplementary Material

Acknowledgments

This work was supported past the Indo–French Centre for the Promotion of Avant-garde Research (project No. 4003-2).

References

• Aloy, P., Pichaud, M. & Russell, R. B. (2005). Curr. Opin. Struct. Biol. 15, 15–22. [PubMed]
• Anderson, D. R., Meyers, Grand. J., Kurumbail, R. G., Caspers, N., Poda, G. I., Long, South. A., Pierce, B. S., Mahoney, 1000. W., Mourey, R. J. & Parikh, 1000. D. (2009). Bioorg. Med. Chem. Lett. 19, 4882–4884. [PubMed]
• Banfield, Yard. J., Naylor, R. L., Robertson, A. Chiliad., Allen, S. J., Dawbarn, D. & Brady, R. Fifty. (2001). Structure, ix, 1191–1199. [PubMed]
• Boehr, D. D., Nussinov, R. & Wright, P. E. (2009). Nat. Chem. Biol. 5, 789–796. [PMC free article] [PubMed]
• Bonvin, A. M. (2006). Curr. Opin. Struct. Biol. 16, 194–200. [PubMed]
• Chakrabarti, P. & Janin, J. (2002). Proteins, 47, 334–343. [PubMed]
• Chakravarty, D., Guharoy, M., Robert, C. H., Chakrabarti, P. & Janin, J. (2013). Protein Sci. 22, 1453–1457. [PMC free article] [PubMed]
• Chen, Fifty., Durley, R. C., Mathews, F. Due south. & Davidson, 5. L. (1994). Science, 264, 86–90. [PubMed]
• Erect, P. J., Antao, T., Chang, J. T., Chapman, B. A., Cox, C. J., Dalke, A., Friedberg, I., Hamelryck, T., Kauff, F., Wilczynski, B. & de Hoon, Thou. J. L. (2009). Bioinformatics, 25, 1422–1423. [PMC gratis commodity] [PubMed]
• Conte, Fifty. L., Chothia, C. & Janin, J. (1999). J. Mol. Biol. 285, 2177–2198. [PubMed]
• Dunker, A. K. et al. (2001). J. Mol. Graph. Model. 19, 26–59. [PubMed]
• Dyson, H. J. & Wright, P. East. (2002). Curr. Opin. Struct. Biol. 12, 54–60. [PubMed]
• Fong, J. H., Shoemaker, B. A., Garbuzynskiy, S. O., Lobanov, M. Y., Galzitskaya, O. V. & Panchenko, A. R. (2009). PLoS Comput. Biol. 5, e1000316. [PMC free article] [PubMed]
• Guharoy, M. & Chakrabarti, P. (2010). BMC Bioinformatics, eleven, 286. [PMC free commodity] [PubMed]
• Guharoy, M., Janin, J. & Robert, C. H. (2010). Proteins, 78, 3219–3225. [PubMed]
• Guharoy, M., Pauwels, Chiliad. & Tompa, P. (2015). Cell, 161, 1230–1230.e1. [PubMed]
• Hillig, R. C., Renault, 50., Vetter, I. R., Drell, T. IV, Wittinghofer, A. & Becker, J. (1999). Mol. Prison cell, 3, 781–791. [PubMed]
• Hubbard, S. J. (1992). NACCESS: A Computer Program for Calculating Accessibilities. Department of Biochemistry and Molecular Biological science, University Higher London.
• Iyer, S., Wei, S., Brew, Thousand. & Acharya, K. R. (2006). J. Biol. Chem. 282, 364–371. [PubMed]
• Janin, J. (2014). Protein Sci. 23, 1813–1817. [PMC gratuitous article] [PubMed]
• Janin, J., Bahadur, R. P. & Chakrabarti, P. (2008). Q. Rev. Biophys. 41, 133–180. [PubMed]
• Jones, S. & Thornton, J. Thou. (1995). Prog. Biophys. Mol. Biol. 63, 31–65. [PubMed]
• Jones, South. & Thornton, J. G. (1997). J. Mol. Biol. 272, 121–132. [PubMed]
• Kabsch, W. & Sander, C. (1983). Biopolymers, 22, 2577–2637. [PubMed]
• Kastritis, P. Fifty., Moal, I. H., Hwang, H., Weng, Z., Bates, P. A., Bonvin, A. M. & Janin, J. (2011). Poly peptide Sci. 20, 482–491. [PMC complimentary article] [PubMed]
• Lee, R. van der et al. (2014). Chem. Rev. 114, 6589–6631. [PMC free article] [PubMed]
• Levy, E. D. (2010). J. Mol. Biol. 403, 660–670. [PubMed]
• Liu, Q., Li, Z. & Li, J. (2014). BMC Bioinformatics, fifteen, Suppl. 16, S3. [PMC complimentary article] [PubMed]
• Liu, R., Jiang, W. & Zhou, Y. (2010). Amino Acids, 38, 263–270. [PubMed]
• Machius, Chiliad., Vértesy, L., Huber, R. & Wiegand, Yard. (1996). J. Mol. Biol. 260, 409–421. [PubMed]
• Maheshwari, S. & Brylinski, M. (2015). J. Mol. Recognit. 28, 35–48. [PubMed]
• Marsh, J. A. & Teichmann, S. A. (2011). Structure, nineteen, 859–867. [PMC free commodity] [PubMed]
• McDonald, I. Thou. & Thornton, J. M. (1994). J. Mol. Biol. 238, 777–793. [PubMed]
• McLachlan, A. D. (1982). Acta Cryst. A38, 871–873.
• Mészáros, B., Tompa, P., Simon, I. & Dosztányi, Z. (2007). J. Mol. Biol. 372, 549–561. [PubMed]
• Mittag, T., Kay, L. E. & Forman-Kay, J. D. (2010). J. Mol. Recognit. 23, 105–116. [PubMed]
• Morgunova, E., Tuuttila, A., Bergmann, U. & Tryggvason, Yard. (2002). Proc. Natl Acad. Sci. USA, 99, 7414–7419. [PMC complimentary commodity] [PubMed]
• Neuvirth, H., Raz, R. & Schreiber, G. (2004). J. Mol. Biol. 338, 181–199. [PubMed]
• Parthasarathy, Due south. & Murthy, Grand. R. Due north. (2000). Poly peptide Eng. Des. Sel. xiii, 9–13.
• Pflugrath, J. W., Wiegand, Chiliad., Huber, R. & Vértesy, 50. (1986). J. Mol. Biol. 189, 383–386. [PubMed]
• Rajamani, D., Thiel, Due south., Vajda, S. & Camacho, C. J. (2004). Proc. Natl Acad. Sci. USA, 101, 11287–11292. [PMC free article] [PubMed]
• Ratnaparkhi, Thou. S., Ramachandran, S., Udgaonkar, J. B. & Varadarajan, R. (1998). Biochemistry, 37, 6958–6966. [PubMed]
• Rice, P., Longden, I. & Bleasby, A. (2000). Trends Genet. 16, 276–277. [PubMed]
• Robinson, R. C., Choe, Due south., Radziejewski, C., Spraggon, G., Greenwald, J., Kostura, M. R., Burtnick, 50. D., Stuart, D. I. & Jones, East. Y. (1999). Protein Sci. viii, 2589–2597. [PMC costless article] [PubMed]
• Ruvinsky, A. M., Kirys, T., Tuzikov, A. V. & Vakser, I. A. (2011). J. Mol. Biol. 408, 356–365. [PMC free article] [PubMed]
• Seewald, M. J., Korner, C., Wittinghofer, A. & Vetter, I. R. (2002). Nature, 415, 662–666. [PubMed]
• Ševčík, J., Urbanikova, 50., Dauter, Z. & Wilson, 1000. S. (1998). Acta Cryst. D54, 954–963. [PubMed]
• Shomura, Y., Dragovic, Z., Chang, H.-C., Tzvetkov, N., Young, J. C., Brodsky, J. L., Guerriero, V., Hartl, F. U. & Bracher, A. (2005). Mol. Prison cell, 17, 367–379. [PubMed]
• Smith, G. R., Sternberg, M. J. & Bates, P. A. (2005). J. Mol. Biol. 347, 1077–1101. [PubMed]
• Stites, W. E. (1997). Chem. Rev. 97, 1233–1250. [PubMed]
• Swapna, Fifty. South., Mahajan, Southward., de Brevern, A. G. & Srinivasan, N. (2012). BMC Struct. Biol. 12, 6. [PMC free article] [PubMed]
• Tompa, P. & Fuxreiter, Yard. (2008). Trends Biochem. Sci. 33, two–8. [PubMed]
• Tuuttila, A., Morgunova, E., Bergmann, U., Lindqvist, Y., Maskos, Chiliad., Fernandez-Catalan, C., Bode, West., Tryggvason, K. & Schneider, Yard. (1998). J. Mol. Biol. 284, 1133–1140. [PubMed]
• White, A., Pargellis, C. A., Studts, J. 1000., Werneburg, B. Thousand. & Farmer, B. T. Two (2007). Proc. Natl Acad. Sci. United states of america, 104, 6353–6358. [PMC free commodity] [PubMed]
• Wiegand, 1000., Epp, O. & Huber, R. (1995). J. Mol. Biol. 247, 99–110. [PubMed]
• Wu, B., Arumugam, S., Gao, Thousand., Lee, Chiliad.-I., Semenchenko, V., Huang, W., Brew, K. & Van Doren, S. R. (2000). J. Mol. Biol. 295, 257–268. [PubMed]
• Zhu, Z., Cunane, L. M., Chen, Z., Durley, R. C., Mathews, F. S. & Davidson, V. L. (1998). Biochemistry, 37, 17128–17136. [PubMed]

---

Articles from IUCrJ are provided hither courtesy of International Union of Crystallography

---

Source: https://www.ncbi.nlm.nih.gov/pmc/articles/PMC4645109/

Posted by: meachamhiscon.blogspot.com

Share this post