Supertertiary protein structure affects an allosteric network

Abstract

Corresponding authors: Stefano.Gianni@uniroma1.it; Per.Jemth@imbim.uu.seThe notion that protein function is allosterically regulated by structural or dynamic changes in proteins has been extensively investigated in several protein domains in isolation. In particular, PDZ domains have represented a paradigm for these studies, despite providing conflicting results. Furthermore, it is still unknown how the association between protein domains in supramodules, consitituting so-called supertertiary structure, affects allosteric networks. Here, we experimentally mapped the allosteric network in a PDZ:ligand complex, both in isolation and in the context of a supramodular structure, and show that allosteric networks in a PDZ domain are highly dependent on the supertertiary structure in which they are present. This striking sensitivity of allosteric networks to presence of adjacent protein domains is likely a common property of supertertiary structures in proteins.Our findings have general implications for prediction of allosteric networks from primary and tertiary structure and for quantitative descriptions of allostery. Abstract IntroductionAllosteric regulation is an essential function of many proteins, well established in multimeric proteins with well-defined conformational changes. From a biological perspective, such allostery plays important roles, for example in regulation of enzyme activity and binding of oxygen to hemoglobin, which was the basis for models such as MWC (1) and KNF (2). More recently the concept has been extended to monomeric proteins (3) and intrinsically disordered proteins (4, 5). Here, allostery is a process, where a signal propagates from one site to a physically distinct site, although the exact mechanism is elusive (6). Thus, as experimental approaches have developed the definition of the allosteric mechanism has evolved too and it is now spanning from the classical structure based allostery to the ensemble nature of allostery, and provides the framework for capturing allostery from rigid and structured proteins to dynamic and disordered proteins (7).PDZ3 from PSD-95 has been extensively used as a model system for allostery in a single protein domain. Originally, it was used as a benchmark of a statistical method to predict allostery from co-variation of mutations (8) and has since then been subject to study by numerous methods (9). PDZ domains are small protein domains, around 100 amino acid residues, with a specific fold that contains 5-6 b strands and 2-3 a-helices ( Figure 1C). The PDZ domain family is one of the most abundant protein-protein interaction domains in humans and it is often found in scaffolding or signalling proteins. PDZ domains bind to short interaction motifs (3-6 residues), usually at the C-terminus of ligand proteins. Looking at PDZ3 as a system to understand principles that underlie allosteric regulation reveals the complexity of the phenomenon. The first allosteric network in PDZ3 was determined by statistical coupling analysis and depicted a network that propagates f...

Global mapping of the energetic and allosteric landscapes of protein binding domains

“…We find excellent agreement between inferred free energies of folding relative to the wild-type (ΔΔG f ) and those corresponding to single AA substitutions determined in vitro for PSD95-PDZ3 (F337W background) 42 (Figure 2e, Pearson’s R = 0.79, n = 30, p-value = 2e-7). Binding free energy changes similarly agree with those measured by stopped-flow experiments for the PSD95-PDZ3:CRIPT interaction 43 (Figure 2e, Pearson’s R = 0.91, n = 26, p-value = 8e-11; see Figure S6 for additional comparisons to smaller-scale in vitro validation datasets). Using only the binding or only single AA substitutions data results in worse agreement with the in vitro binding and folding free energy changes (Figure S4), as does reducing the number of double mutants used to fit the model (Figure S5).…”

Multisite NHERF1 phosphorylation controls GRK6A regulation of hormone-sensitive phosphate transport

“…7), which is required for inhibition of phosphate transport (16). Allosteric networks and allosteric communication are essential regulators of PDZ-containing multidomain protein conformational changes and function (44,45).…”

Evotuning protocols for Transformer-based variant effect prediction on multi-domain proteins

“…Next, for the category C proteins with cooperative domains, the protocol aiming to gather evolutionary information from sequences covering inter-domain interactions (i.e., protocol C) realized the best results. In a rare case (PSD95 with training size 0.8), the performance of the protocol B was comparable to the protocol C. A possible reason is that the mutagenesis assay was conducted for the isolated PDZ domain, not for the full length [40]. Nonetheless, in the remaining cases, the protocol C was better than the protocol B.…”

“…Here we classified the proteins into four categories considering their domain architecture and (non-)existence of intramolecular domain interactions. The single-domain proteins (TEM-1 [11] and APH(3’)-II [12]) were classified into the category A; in the category B, the multi-domain proteins without intramolecular domain interactions were collected (ubiquitin [13], YAP65 [14], and E3 ligase [15]); the proteins known to have interacting domains fell into the category C (PSD95 [16, 40] and Pab1 [17,41,42]). Each of these protein categories corresponded to the respective protein engineering scenarios described above.…”