Danger in the dust.

Background: Allosteric communications are vital for cellular signaling. Here we explore a relationship between protein architectural organization and shortcuts in signaling pathways. Results: We show that protein domains consist of modules interconnected by residues that mediate signaling through the shortest pathways. These mediating residues tend to be located at the inter-modular boundaries, which are more rigid and display a larger number of long-range interactions than intra-modular regions. The inter-modular boundaries contain most of the residues centrally conserved in the protein fold, which may be crucial for information transfer between amino acids. Our approach to modular decomposition relies on a representation of protein structures as residue-interacting networks, and removal of the most central residue contacts, which are assumed to be crucial for allosteric communications. The modular decomposition of 100 multi-domain protein structures indicates that modules constitute the building blocks of domains. The analysis of 13 allosteric proteins revealed that modules characterize experimentally identified functional regions. Based on the study of an additional functionally annotated dataset of 115 proteins, we propose that high-modularity modules include functional sites and are the basic functional units. We provide examples (the Gαs subunit and P450 cytochromes) to illustrate that the modular architecture of active sites is linked to their functional specialization. Conclusion: Our method decomposes protein structures into modules, allowing the study of signal transmission between functional sites. A modular configuration might be advantageous: it allows signaling proteins to expand their regulatory linkages and may elicit a broader range of control mechanisms either via modular combinations or through modulation of inter-modular linkages. Published: 25 May 2007 Genome Biology 2007, 8:R92 (doi:10.1186/gb-2007-8-5-r92) Received: 10 October 2006 Revised: 6 February 2007 Accepted: 25 May 2007 The electronic version of this article is the complete one and can be found online at http://genomebiology.com/2007/8/5/R92 Genome Biology 2007, 8:R92 R92.2 Genome Biology 2007, Volume 8, Issue 5, Article R92 del Sol et al. http://genomebiology.com/2007/8/5/R92 Background Allosteric communications play crucial roles in many cellular signaling processes. Perturbations caused by factors such as ligand binding at one functional site affect a distant site, thereby regulating binding affinity and catalytic activity [1,2]. Since the allosteric model proposed by Monod and coworkers [1], decades of research have extended the common view of allostery associated with multi-domain proteins to single domain proteins. The allosteric behavior displayed by single domain proteins, such as myoglobin [3], called into question the existing allosteric dogma. In the 'new view' of protein allostery, all proteins are potentially allosteric when thought of in terms of population redistribution upon ligand binding causing conformational change in a second binding site [1]. Dynamic models have been proposed to explain the conformational changes involved in signal transmission between functional sites [4,5]. In particular, the role of the pre-existing equilibrium of conformational sub-states in allostery proposed already over 20 years ago [6] is increasingly receiving attention, emphasizing the key role of protein dynamics in this process [1,7-9]. Although experimental methods such as double mutant cycle analysis [10] have provided insights into allosteric communications, understanding the general principles of the transmission of information between distant functional surfaces remains a challenge in structural biology. Several theoretical methods based on sequence and structural considerations have been proposed for the identification of key amino acids for long-range communications [11-13]. Among these, an interesting sequence-based approach has been proposed by Ranganathan and coworkers [14,15] for estimating the thermodynamic coupling between amino acids in several examples of protein families. Recently, we introduced a model based on a network representation of protein structures. The model allows us to determine fold centrally conserved residues (FCCRs). These residues are responsible for maintaining the shortest pathways between all amino acids and, thus, play key roles in signal transmission [13]. Analysis of several protein families showed an agreement between our results and experimental data, illustrating the importance of protein topology in network communications. Perceiving protein structures as information processing networks, it is reasonable to assume that mutations of amino acids crucial for network communications could impair signal transmission. The rationale for modular organization of proteins in allosteric behavior has been discussed previously [16-18]. Modular domains can act cooperatively, leading to new input (and output) relationships. The Src family proteins constitute a clear example of this modular architecture: these proteins contain amino-terminal SH3 and SH2 domains, which flank a kinase domain by intra-molecular SH3-binding and SH2binding sites [16]. It is further known that modular functional units display certain degrees of functional specificity in a number of proteins. In several cases of protein-protein interactions, which are involved in cell signaling, some parts of the interacting interface participate in the information transfer, whereas other interacting regions appear to contribute solely to binding affinity [19]. Examples of proteins exhibiting this binding site modular configuration include Myosin, C5a receptor, and the protein kinase R activator PACT among others [19]. Here, we aim to obtain the modular decomposition of allosteric proteins and to explore a relationship between the modules and the allosteric activity. We expect that such a relationship, if it exists, would lead to deeper insight into functional mechanisms. We develop a new approach for decomposing protein structures into modules using their residue network representations. Our methodology is based on the edge-betweenness clustering algorithm proposed by Newman and Girvan [20,21], which has been previously applied to a wide variety of problems [22-25]. This method uses edge centrality to detect module boundaries and finds the assignation of nodes into modules [20]. The small-world topology of protein structures suggests that the key amino acids for signal transmission should lie in the shortcuts linking different regions of the structure. The removal of the most central contacts forming these shortcuts divides the structure into modules. We characterize these modules from a structural point of view. Our results, derived from a non-redundant dataset of multi-domain proteins, reveal that, in the vast majority of the cases, modules tend to be located within rather than across domains. Therefore, modules can be considered as sub-domains. Further analysis shows that the percentage of long-range interactions at the modular boundaries is much higher than that in non-boundary regions. Residues forming inter-modular contacts fluctuate less than those participating only in the intra-modular interactions. One possible explanation of this finding is that most central residues, which have been shown to be important for the allosteric communications, are located at the inter-modular interfaces and, therefore, tend to be more rigid to maintain their contacts. Inspection of 13 allosteric proteins shows that functionally annotated regions exhibit a modular architecture, with modules interconnected by FCCRs, which are responsible for mediating the shortest pathways between all amino acids and, thus, play crucial roles in allosteric communications [13]. Functional sites are often contained in one module; however, there are also examples of functional sites shared by two or more modules. Some of these cases correspond to binding sites divided into two modules belonging to different domains. The Gαs subunit and P450 cytochromes are examples of functional sites shared between modules. Interestingly, the modular decomposition of the Gαs subunit reflects binding site partitioning into regions involved in different sub-functional specialization, general binding and information transfer regions [26]. The P450eryF active site is divided into a module containing the ligand-binding site, and a module comprising the effectorbinding site, whereas the P450cam substrate binds to one module, and the product binds mainly to another module. A Genome Biology 2007, 8:R92 http://genomebiology.com/2007/8/5/R92 Genome Biology 2007, Volume 8, Issue 5, Article R92 del Sol et al. R92.3