Characterization of Protein–Protein Interfaces

We analyze the characteristics of protein–protein interfaces using the largest datasets available from the Protein Data Bank (PDB). We start with a comparison of interfaces with protein cores and non-interface surfaces. The results show that interfaces differ from protein cores and non-interface surfaces in residue composition, sequence entropy, and secondary structure. Since interfaces, protein cores, and non-interface surfaces have different solvent accessibilities, it is important to investigate whether the observed differences are due to the differences in solvent accessibility or differences in functionality. We separate out the effect of solvent accessibility by comparing interfaces with a set of residues having the same solvent accessibility as the interfaces. This strategy reveals residue distribution propensities that are not observable by comparing interfaces with protein cores and non-interface surfaces. Our conclusions are that there are larger numbers of hydrophobic residues, particularly aromatic residues, in interfaces, and the interactions apparently favored in interfaces include the opposite charge pairs and hydrophobic pairs. Surprisingly, Pro-Trp pairs are over represented in interfaces, presumably because of favorable geometries. The analysis is repeated using three datasets having different constraints on sequence similarity and structure quality. Consistent results are obtained across these datasets. We have also investigated separately the characteristics of heteromeric interfaces and homomeric interfaces.     

Predicting Protein–Protein Interfaces that Bind Intrinsically Disordered Protein Regions

A long-standing goal in biology is the complete annotation of function and structure on all protein–protein interactions, a large fraction of which is mediated by intrinsically disordered protein regions (IDRs). However, knowledge derived from experimental structures of such protein complexes is disproportionately small due, in part, to challenges in studying interactions of IDRs. Here, we introduce IDRBind, a computational method that by combining gradient boosted trees and conditional random field models predicts binding sites of IDRs with performance approaching state-of-the-art globular interface predictions, making it suitable for proteome-wide applications. Although designed and trained with a focus on molecular recognition features, which are long interaction-mediating-elements in IDRs, IDRBind also predicts the binding sites of short peptides more accurately than existing specialized predictors. Consistent with IDRBind's specificity, a comparison of protein interface categories uncovered uniform trends in multiple physicochemical properties, positioning molecular recognition feature interfaces between peptide and globular interfaces.     

Keep Your Fingers Off My DNA: Protein–Protein Interactions Mediated by C2H2 Zinc Finger Domains

Cys2-His2 (C2H2) zinc finger domains (ZFs) were originally identified as DNA-binding domains, and uncharacterized domains are typically assumed to function in DNA binding. However, a growing body of evidence suggests an important and widespread role for these domains in protein binding. There are even examples of zinc fingers that support both DNA and protein interactions, which can be found in well-known DNA-binding proteins such as Sp1, Zif268, and Ying Yang 1 (YY1). C2H2 protein–protein interactions (PPIs) are proving to be more abundant than previously appreciated, more plastic than their DNA-binding counterparts, and more variable and complex in their interactions surfaces. Here we review the current knowledge of over 100 C2H2 zinc finger-mediated PPIs, focusing on what is known about the binding surface, contributions of individual fingers to the interaction, and function. An accurate understanding of zinc finger biology will likely require greater insights into the potential protein interaction capabilities of C2H2 ZFs.     

Protein–Protein and Protein–Membrane Associations in the Lignin Pathway

Supramolecular organization of enzymes is proposed to orchestrate metabolic complexity and help channel intermediates in different pathways. Phenylpropanoid metabolism has to direct up to 30% of the carbon fixed by plants to the biosynthesis of lignin precursors. Effective coupling of the enzymes in the pathway thus seems to be required. Subcellular localization, mobility, protein–protein, and protein–membrane interactions of four consecutive enzymes around the main branch point leading to lignin precursors was investigated in leaf tissues of Nicotiana benthamiana and cells of Arabidopsis thaliana. CYP73A5 and CYP98A3, the two Arabidopsis cytochrome P450s (P450s) catalyzing para- and meta-hydroxylations of the phenolic ring of monolignols were found to colocalize in the endoplasmic reticulum (ER) and to form homo- and heteromers. They moved along with the fast remodeling plant ER, but their lateral diffusion on the ER surface was restricted, likely due to association with other ER proteins. The connecting soluble enzyme hydroxycinnamoyltransferase (HCT), was found partially associated with the ER. Both HCT and the 4-coumaroyl-CoA ligase relocalized closer to the membrane upon P450 expression. Fluorescence lifetime imaging microscopy supports P450 colocalization and interaction with the soluble proteins, enhanced by the expression of the partner proteins. Protein relocalization was further enhanced in tissues undergoing wound repair. CYP98A3 was the most effective in driving protein association.     

Computational Prediction of Protein–Protein Interactions

Recently a number of computational approaches have been developed for the prediction of protein–protein interactions. Complete genome sequencing projects have provided the vast amount of information needed for these analyses. These methods utilize the structural, genomic, and biological context of proteins and genes in complete genomes to predict protein interaction networks and functional linkages between proteins. Given that experimental techniques remain expensive, time-consuming, and labor-intensive, these methods represent an important advance in proteomics. Some of these approaches utilize sequence data alone to predict interactions, while others combine multiple computational and experimental datasets to accurately build protein interaction maps for complete genomes. These methods represent a complementary approach to current high-throughput projects whose aim is to delineate protein interaction maps in complete genomes. We will describe a number of computational protocols for protein interaction prediction based on the structural, genomic, and biological context of proteins in complete genomes, and detail methods for protein interaction network visualization and analysis.     

Targeting Protein for Xenopus Kinesin-like Protein 2 (TPX2) Regulates γ-Histone 2AX (γ-H2AX) Levels upon Ionizing Radiation

The microtubule-associated protein targeting protein for Xenopus kinesin-like protein 2 (TPX2) plays a key role in spindle assembly and is required for mitosis in human cells. In interphase, TPX2 is actively imported into the nucleus to prevent its premature activity in microtubule organization. To date, no function has been assigned to nuclear TPX2. We now report that TPX2 plays a role in the cellular response to DNA double strand breaks induced by ionizing radiation. Loss of TPX2 leads to inordinately strong and transient accumulation of ionizing radiation-dependent Ser-139-phosphorylated Histone 2AX (γ-H2AX) at G₀ and G₁ phases of the cell cycle. This is accompanied by the formation of increased numbers of high intensity γ-H2AX ionizing radiation-induced foci. Conversely, cells overexpressing TPX2 have reduced levels of γ-H2AX after ionizing radiation. Consistent with a role for TPX2 in the DNA damage response, we found that the protein accumulates at DNA double strand breaks and associates with the mediator of DNA damage checkpoint 1 (MDC1) and the ataxia telangiectasia mutated (ATM) kinase, both key regulators of γ-H2AX amplification. Pharmacologic inhibition or depletion of ATM or MDC1, but not of DNA-dependent protein kinase (DNA-PK), antagonizes the γ-H2AX phenotype caused by TPX2 depletion. Importantly, the regulation of γ-H2AX signals by TPX2 is not associated with apoptosis or the mitotic functions of TPX2. In sum, our study identifies a novel and the first nuclear function for TPX2 in the cellular responses to DNA damage.     

Protein Phosphatase 2A Isoforms Utilizing Aβ Scaffolds Regulate Differentiation through Control of Akt Protein

Protein phosphatase 2A (PP2A) regulates almost all cell signaling pathways. It consists of a scaffolding A subunit to which a catalytic C subunit and one of many regulatory B subunits bind. Of the more than 80 PP2A isoforms, 10% use Aβ as a scaffold. This study demonstrates the isoform-specific function of the A scaffold subunits. Polyomaviruses have shown the importance of phosphotyrosine, PI3K, and p53 in transformation. Comparisons of polyoma and SV40 small T antigens implicate Aβ in the control of differentiation. Knockdown of Aβ enhanced differentiation. Akt signaling regulated differentiation; its activation or inhibition promoted or blocked it, respectively. Aβ bound Akt. Enhancement of PP2A Aβ/Akt interaction by polyoma small T antigen increased turnover of Akt Ser-473 phosphorylation. Conversely, knockdown of Aβ promoted Akt activity and reduced turnover of phosphate at Ser-473 of Akt. These data provide new insight into the regulation of Akt, a protein of extreme importance in cancer. Furthermore, our results suggest that the role for Aβ in differentiation and perhaps tumor suppression may lie partly in its ability to negatively regulate Akt.     

Protein phosphatase 2A--a 'ménage à trois'

Protein phosphorylation is probably the major regulatory mechanism employed by eukaryotic cells. Much work has been devoted to the role of protein kinases and their modulation by hormones, growth factors and neurotransmitters. It is now appreciated that protein phosphatases are also key players in actively regulating many cellular processes. In this article we discuss recent advances in our understanding of the function of protein phosphatase 2A, one of the major serine/threonine-specific protein phosphatases.     

Protein Phosphatase 6 Interacts with the DNA-Dependent Protein Kinase Catalytic Subunit and Dephosphorylates γ-H2AX

The catalytic subunit of the DNA-dependent protein kinase (DNA-PKcs) plays a major role in the repair of DNA double-strand breaks (DSBs) by nonhomologous end joining (NHEJ). We have previously shown that DNA-PKcs is autophosphorylated in response to ionizing radiation (IR) and that dephosphorylation by a protein phosphatase 2A (PP2A)-like protein phosphatase (PP2A, PP4, or PP6) regulates the protein kinase activity of DNA-PKcs. Here we report that DNA-PKcs interacts with the catalytic subunits of PP6 (PP6c) and PP2A (PP2Ac), as well as with the PP6 regulatory subunits PP6R1, PP6R2, and PP6R3. Consistent with a role in the DNA damage response, silencing of PP6c by small interfering RNA (siRNA) induced sensitivity to IR and delayed release from the G₂/M checkpoint. Furthermore, siRNA silencing of either PP6c or PP6R1 led to sustained phosphorylation of histone H2AX on serine 139 (γ-H2AX) after IR. In contrast, silencing of PP6c did not affect the autophosphorylation of DNA-PKcs on serine 2056 or that of the ataxia-telangiectasia mutated (ATM) protein on serine 1981. We propose that a novel function of DNA-PKcs is to recruit PP6 to sites of DNA damage and that PP6 contributes to the dephosphorylation of γ-H2AX, the dissolution of IR-induced foci, and release from the G₂/M checkpoint in vivo.DNA double-strand breaks (DSBs) are the most cytotoxic form of DNA damage. In human cells there are two main pathways for the repair of DSBs, namely, nonhomologous end joining (NHEJ) and homologous recombination (HR) (reviewed in reference 26). In the initial phase of NHEJ, DSBs are detected by the Ku70/80 heterodimer, which leads to recruitment of the DNA-dependent protein kinase catalytic subunit (DNA-PKcs) and stimulation of its serine/threonine protein kinase activity. Upon autophosphorylation, DNA-PKcs undergoes a conformational change and dissociates from the DSB (25), providing other DNA repair proteins with access to the damage site (reviewed in reference 33). Another physiological substrate of DNA-PK is a histone H2A variant, H2AX. DNA-PKcs and the related protein kinase ATM (ataxia-telangiectasia mutated) both contribute to DNA damage-induced phosphorylation of H2AX on serine 139 to form γ-H2AX (51), which acts as a recruitment platform for MDC1, 53BP1, and other proteins involved in the DNA damage response and cell cycle checkpoint activation (7, 52).While the effects of phosphorylation on the repair process have been well documented, comparatively little is known about the role of serine/threonine phosphoprotein phosphatases (PPPs) in the DNA damage response. Within the PPP family, the catalytic subunits of PP2A (PP2Ac), PP4 (PP4c), and PP6 (PP6c) are most closely related and form a subgroup referred to as the PP2A-like protein phosphatases (reviewed in reference 40). In vitro, the PP2A-like enzymes display similar sensitivities to small-molecule inhibitors such as okadaic acid and microcystin (27, 45, 53). The specificity of PP2Ac, PP4c, and PP6c function in vivo is derived from a group of regulatory subunits that, with the exception of α4/TAP42 and TIP41, are unique to each enzyme (12, 13, 27, 45, 49). PP2Ac associates with a scaffolding A-α or A-β subunit and additional B-type subunits, while four direct binding partners and several other complex partners unique to PP4c have been characterized (12). The Saccharomyces cerevisiae homologue of PP6c, known as Sit4, interacts with three related proteins: the Sit4-associated proteins SAP155, SAP185, and SAP190, each of which contains a conserved domain known as the SAPs domain (32, 50). The SAPs domain is present in three human orthologues designated PP6R1, PP6R2, and PP6R3, which are therefore considered PP6c regulatory subunits, and each has been shown to bind independently to PP6c (48). More recently, three ankyrin repeat-containing proteins (ARS-A, ARS-B, and ARS-C) were identified as PP6R1 binding partners. One of these, ARS-A, has been shown to dock all three SAPs domain proteins (50), suggesting that, like PP2Ac, PP6c forms stable heterotrimers in vivo and that together these subunits define PP6 function.We have previously shown that inhibition of PP2A-like protein phosphatase activity by okadaic acid increases the phosphorylation status of DNA-PKcs and decreases its protein kinase activity (20), thus implicating PP2A-like phosphatases in the regulation of DNA-PK activity in vivo. More recently, both PP4 and PP2A have been shown to play roles in the DNA damage response by dephosphorylating γ-H2AX (14, 15, 28, 42). However, the potential role of PP6 in γ-H2AX dephosphorylation has not been addressed.Here we show that DNA-PKcs interacts with PP2Ac and PP6c, as well as with the PP6c regulatory subunits, PP6R1, PP6R2, and PP6R3. Depletion of PP6c by small interfering RNA (siRNA) induces sensitivity to ionizing radiation (IR) and delayed release from the G₂/M checkpoint. Furthermore, siRNA silencing of either PP6c or PP6R1 leads to sustained phosphorylation of γ-H2AX after DNA damage. Together, our studies reveal that a novel and previously unrecognized function of DNA-PKcs may be to recruit PP6 to sites of DNA damage and that PP6 regulates the phosphorylation status of γ-H2AX, the dissolution of IR-induced foci, and release from the G₂/M checkpoint.     

Peptide, Peptidomimetic, and Small-molecule Antagonists of the p53–HDM2 Protein–Protein Interaction

Modulation of intracellular protein–protein interactions has been – and remains – a challenging goal for the discovery and development of small-molecule therapeutic agents. Progress in the pharmacological targeting and understanding at the molecular level of one such interaction that is relevant to cancer drug research, viz. that between the tumour suppressor protein p53 and its negative regulator HDM2, is reviewed here. The first X-ray crystal structure of a complex between a small peptide from the trans-activation domain of p53 and the N-terminal domain of HDM2 was reported almost 10 years ago. The nature of this interaction, which involves just three residue side chains in the p53 peptide ligand and a compact hydrophobic binding pocket in the HDM2 receptor, together with the attractive concept of reactivating the anti-proliferative functions of p53 in tumour cells, has spurned a great deal of effort aimed at finding drug-like antagonists of this interaction. A variety of approaches, including both structure-guided peptidomimetic and de novo design, as well as high through-put screening campaigns, have provided a wealth of leads that might be turned into actual drugs. There is still some way to go as far as optimisation and preclinical development of such leads is concerned, but it is clear already now that antagonists of the p53–HDM2 protein–protein interaction have a good chance of ultimately being successful in providing a new anti-cancer therapy modality, both in monotherapy and to potentiate the effectiveness of existing chemotherapies.     

β-Arrestin 2 Promotes Hepatocyte Apoptosis by Inhibiting Akt Protein

Recent studies reveal that multifunctional protein β-arrestin 2 (Arrb2) modulates cell apoptosis. Survival and various aspects of liver injury were investigated in WT and Arrb2 KO mice after bile duct ligation (BDL). We found that deficiency of Arrb2 enhances survival and attenuates hepatic injury and fibrosis. Following BDL, Arrb2-deficient mice as compared with WT controls displayed a significant reduction of hepatocyte apoptosis as demonstrated by the TUNEL assay. Following BDL, the levels of phospho-Akt and phospho-glycogen synthase kinase 3β (GSK3β) in the livers were significantly increased in Arrb2 KO compared with WT mice, although p-p38 increased in WT but not in Arrb2-deficient mice. Inhibition of GSK3β following BDL decreases hepatic apoptosis and decreased p-p38 in WT mice but not in Arrb2 KO mice. Activation of Fas receptor with Jo2 reduces phospho-Akt and increases apoptosis in WT cells and WT mice but not in Arrb2-deficient cells and Arrb2-deficient mice. Consistent with direct interaction of Arrb2 with and regulating Akt phosphorylation, the expression of a full-length or N terminus but not the C terminus of Arrb2 reduces Akt phosphorylation and coimmunoprecipates with Akt. These results reveal that the protective effect of deficiency of Arrb2 is due to loss of negative regulation of Akt due to BDL and decreased downstream GSK3β and p38 MAPK signaling pathways.     

Protein–ligand interaction-guided discovery of novel VEGFR-2 inhibitors

Abstract

As an effective target in abnormal angiogenesis-related tumor treatment, VEGFR-2 has small-molecule inhibitors of various scaffolds being approved for treating diseases such as renal carcinoma, non-small cell lung cancer, etc. However, endogenous and acquired drug resistance are still considered to be the main contributors for the failure of VEGFR-2 clinical candidates. Therefore, development of novel VEGFR-2 inhibitors is still urgently needed in the market but also challenging. In this work, residues including Asp1046, Ile1025, HIS1026, Cys919 and Lys868 were identified as the most important residues for Hbonded interaction, while His1026, Asp1046, Glu885, Ile1025 and Leu840 exhibited critical role for the nonbonded interactions through a comprehensive analysis of protein–ligand interactions, which plays critical roles in the binding of compounds and targets. Guided by the analysis of binding interactions, a total of 10 novel VEGFR-2 inhibitors based on N-methyl-4-oxo-N-propyl-1,4-dihydroquinoline-2-carboxamide scaffold were discovered through fragment-based drug design and structure-based virtual screening, which expands the chemical space of current VEGFR-2 inhibitors. Biological activity evaluation showed that even though the enzymatic activity of these compounds against VEGFR-2 were inferior to that of the positive controls sorafenib and motesanib, compound I-10 showed moderate HepG2 cell inhibitory activity with an IC₅₀ value of 33.65?μM and eight compounds exhibited moderate or higher HUVEC inhibitory activity in the range of 19.54–57.98?μM compared to the controls. Particularly, the HUVEC inhibitory activity of compound I-6 (IC₅₀ = 19.54?μM) outperformed motesanib and can be used as starting points for further optimization and development for cancer treatment.

Communicated by Ramaswamy H. Sarma     

Protein–Protein Interactions in DNA Base Excision Repair

The system of base excision repair (BER) ensures correction of the most abundant DNA damages in mammalian cells and plays an important role in maintaining genome stability. Enzymes and protein factors participate in the multistage BER in a coordinated fashion, which ensures repair efficiency. The suggested coordination mechanisms are based on formation of protein complexes stabilized via either direct or indirect DNA-mediated interactions. The results of investigation of direct interactions of the proteins participating in BER with each other and with other proteins are outlined in this review. The known protein partners and sites responsible for their interaction are presented for the main participants as well as quantitative characteristics of their affinity. Information on the mechanisms of regulation of protein–protein interactions mediated by DNA intermediates and posttranslational modification is presented. It can be suggested based on all available data that the multiprotein complexes are formed on chromatin independent of the DNA damage with the help of key regulators of the BER process – scaffold protein XRCC1 and poly(ADP-ribose) polymerase 1. The composition of multiprotein complexes changes dynamically depending on the DNA damage and the stage of BER process.     

Rapid Production of Platelet-activating Factor Is Induced by Protein Kinase Cα-mediated Phosphorylation of Lysophosphatidylcholine Acyltransferase 2 Protein

Platelet-activating factor (PAF), a potent proinflammatory lipid mediator, is synthesized rapidly in response to extracellular stimuli by the activation of acetyl-CoA:lyso-PAF acetyltransferase (lyso-PAFAT). We have reported previously that lyso-PAFAT activity is enhanced in three distinct ways in mouse macrophages: rapid activation (30 s) after PAF stimulation and minutes to hours after LPS stimulation. Lysophosphatidylcholine acyltransferase 2 (LPCAT2) was later identified as a Ca²⁺-dependent lyso-PAFAT. However, the mechanism of rapid lyso-PAFAT activation within 30 s has not been elucidated. Here we show a new signaling pathway for rapid biosynthesis of PAF that is mediated by phosphorylation of LPCAT2 at Ser-34. Stimulation by either PAF or ATP resulted in PKCα-mediated phosphorylation of LPCAT2 to enhance lyso-PAFAT activity and rapid PAF production. Biochemical analyses showed that the phosphorylation of Ser-34 resulted in augmentation of V_max with minimal K_m change. Our results offer an answer for the previously unknown mechanism of rapid PAF production.     

A Critical Assessment of Information-guided Protein–Protein Docking Predictions

The structures of protein complexes are increasingly predicted via protein–protein docking (PPD) using ambiguous interaction data to help guide the docking. These data often are incomplete and contain errors and therefore could lead to incorrect docking predictions. In this study, we performed a series of PPD simulations to examine the effects of incompletely and incorrectly assigned interface residues on the success rate of PPD predictions. The results for a widely used PPD benchmark dataset obtained using a new interface information-driven PPD (IPPD) method developed in this work showed that the success rate for an acceptable top-ranked model varied, depending on the information content used, from as high as 95% when contact relationships (though not contact distances) were known for all residues to 78% when only the interface/non-interface state of the residues was known. However, the success rates decreased rapidly to ∼40% when the interface/non-interface state of 20% of the residues was assigned incorrectly, and to less than 5% for a 40% incorrect assignment. Comparisons with results obtained by re-ranking a global search and with those reported for other data-guided PPD methods showed that, in general, IPPD performed better than re-ranking when the information used was more complete and more accurate, but worse when it was not, and that when using bioinformatics-predicted information on interface residues, IPPD and other data-guided PPD methods performed poorly, at a level similar to simulations with a 40% incorrect assignment. These results provide guidelines for using information about interface residues to improve PPD predictions and reveal a bottleneck for such improvement imposed by the low accuracy of current bioinformatic interface residue predictions.Proteins work in close association with other proteins to mediate the intricate functions of a cell. The atomic resolution of the structure of a protein complex can therefore help one understand a protein''s function in detail. Protein–protein docking (PPD),¹ a computational approach that complements experimental structure determinations, has attracted increasing research interest (1, 2), in part because it remains challenging to determine most structures of protein complexes via experimental techniques (3).To improve the performance of PPD predictions, experimentally derived data (e.g. distances) and information (e.g. the identity of interface residues) have been used either as a filter allowing less plausible docking solutions to be disregarded (4–9) or as a constraint to guide the docking process (10, 11). Various types of data and information have been used to aid PPD (12); these range from distances between, or the relative orientation of, the two interacting proteins to simple identification of the amino acid residues directly involved in the binding of the two proteins (13). Despite considerable success, the caveat for all these data-guided PPD predictions is that the data or information used must be correct in order to avoid spurious results caused by misguiding (12). It is therefore pertinent and important to evaluate the effects of errors in the incorporated data or information on the quality of PPD solutions.We have recently shown that the use of just a few distance constraints can improve the success rates of PPD such that they rival, or are even better than, those of a global search ranked using a sophisticated energy function, and that errors in the distance data significantly decrease the success rates of prediction (11). However, because distance data for interacting proteins are usually hard to obtain, other types of data or information, even if “ambiguous” (10), are increasingly used in PPD predictions (12, 14). In this study, we investigated the effects of incompletely and incorrectly assigned interface/non-interface residues, a major source of the so-called ambiguous data, on information-guided PPD predictions.As illustrated in Fig. 1, the information content of interface/non-interface residues can be rich enough to reveal the identity of every pair of residues in contact, but not their contact distances, or so poor as to reveal the interface/non-interface state of these residues but not their pairing relationship, for one or both of the two interacting proteins. To determine how these different levels of residue information content can help PPD predictions and the extent to which the use of incorrectly assigned residues degrades prediction success rates, we have developed a new interface information-driven PPD method (IPPD) and carried out a series of PPD simulations on a well-tested benchmark dataset. The results showed that when the information content was rich, excellent predictions (success rates for producing an acceptable top-ranked model > 70%) could be made via IPPD or by re-ranking a global search''s solutions using the same interface information, and that, encouragingly, the success of predictions remained respectable (top-ranked success rates > 15%) when the content was poor. However, when enough of the interface residues were incorrectly assigned, as would be the case when using interface residues predicted by a state-of-the-art bioinformatics method such as CPORT (15), few models ranked first by IPPD or other PPD methods, including HADDOCK (10), a popular ambiguous data-driven PPD method, came close to being acceptable. These results suggest that we can greatly increase the power of PPD predictions for practical applications only if the accuracy of current bioinformatics methods for predicting the interface residues of protein complexes can be significantly improved.Open in a separate windowFig. 1.Contact matrix of two interacting proteins, A and B, and the contact vectors of their residues. In the contact matrix, M_ij = 1 or 0, respectively, denotes contact or a lack of contact between residue i in protein A and residue j in protein B. In the contact vectors, V_Ai = 1 or 0, respectively, when residue Ai has, or does not have, at least one contact with any residue of protein B.     

Interaction of Insulin-Like Growth Factor-Binding Protein 2 with α2-Macroglobulin in the Circulation

Insulin-like growth factors (IGFs) play active role in mitogenic and metabolic processes. In the peripheral circulation, they are mostly bound to specific IGF-binding proteins (IGFBPs). Proteolysis of IGFBPs releases free, active IGFs. IGFBP-2 is the second most abundant of the six binding proteins and its concentration increases in catabolic states. The possible interaction between IGFBP-2 and other proteins in the circulation was investigated in this study. Our results showed that IGFBP-2 associates with α2-macroglobulin (α2M), a protease inhibitor. Formation of IGFBP-2/α2M complexes most likely contributes to the regulation of IGFBP-2 proteolysis and, thus, the activity of IGFs.