Thermodynamic consequences of disrupting a water‐mediated hydrogen bond network in a protein:pheromone complex

Sharrow, Scott D.; Edmonds, Katherine A.; Goodman, Michael A.; Novotný, Miloš V.; Stone, Martin J.

doi:10.1110/ps.04912605

Cited by 43 publications

(36 citation statements)

References 42 publications

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“…In this respect, substitutions at position Asn-114 in Abl-SH3 resulted in significant enthalpic effects that correlated very well with the modulation of the watermediated interactions. Even though the enthalpy changes were modest, and were in the lower limit of the values associated to the displacement of water molecules in other systems (28,30), it is important to stress that they only reflect very subtle perturbations in the interactions within one of the hydration clusters, mostly associated with changes in the dynamic properties of waters at hydration site 5. If the observed effects on water occupancy at this position are extrapolated to the 80% occupancy found for the WT-p41 complex, we can tentatively estimate the contribution to the binding enthalpy of water molecules at site 5 to be around Ϫ20 kJ⅐mol Ϫ1 .…”

Section: Discussionmentioning

confidence: 95%

“…A 12-Å cutoff was applied to treat nonbonded interactions, and the particle mesh Ewald method was introduced for the treatment of long range electrostatic interactions. MD trajectories were recorded every 2 ps, analyzed using the ptraj module of AMBER 8.0 (28), and visualized using the VMD program (29). In all cases, the crystallographic water molecules included in the starting structures were replaced in the early stages of the MD simulations by molecules that penetrate spontaneously from the bulk and occupy positions equivalent to those found in the crystal structures.…”

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confidence: 99%

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Role of Interfacial Water Molecules in Proline-rich Ligand Recognition by the Src Homology 3 Domain of Abl

Palencia

Cámara-Artigas

Pisabarro

et al. 2010

Journal of Biological Chemistry

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The interaction of Abl-Src homology 3 domain (SH3) with the high affinity peptide p41 is the most notable example of the inconsistency existing between the currently accepted description of SH3 complexes and their binding thermodynamic signature. We had previously hypothesized that the presence of interfacial water molecules is partially responsible for this thermodynamic behavior. We present here a thermodynamic, structural, and molecular dynamics simulation study of the interaction of p41 with Abl-SH3 and a set of mutants designed to alter the water-mediated interaction network. Our results provide a detailed description of the dynamic properties of the interfacial water molecules and a molecular interpretation of the thermodynamic effects elicited by the mutations in terms of the modulation of the water-mediated hydrogen bond network. In the light of these results, a new dual binding mechanism is proposed that provides a better description of proline-rich ligand recognition by Abl-SH3 and that has important implications for rational design.The recognition of proline-rich sequences by protein-protein interaction modules, such as SH3 3 or WW domains, is one of the most common mechanisms by which specific, transient protein-protein interactions are established within the cell. SH3 domains are found in oncoproteins and proteins overexpressed in deregulated signaling pathways during cancer development and are also associated with other pathologies such as AIDS, osteoporosis, or inflammatory processes (1-2). Inhibitors of the interactions between SH3 domains and their partners have proved to be promising therapeutic agents, validating these domains as attractive targets for drug design (3-6). Nevertheless, despite the wealth of structural and functional information collected during the last 2 decades, the forces driving proline-rich ligand recognition by SH3 domains are still not fully understood.SH3 domains fold into a ␤-barrel structure composed of two orthogonal anti-parallel three-stranded ␤-sheets connected by three main loops (RT, n-Src, and distal loops). The binding site is a relatively flat, hydrophobic surface that consists of three shallow pockets. SH3 ligands typically contain the PpP motif (where and p are frequently hydrophobic and proline residues, respectively) and bind in a PPII conformation so that each of the P moieties packs tightly into one hydrophobic pocket formed by highly conserved aromatic residues on the surface of the domain. Additional interactions, which have been proposed to confer increased affinity and specificity, are established between residues flanking the core motif in the ligand and a third pocket delimited by the RT and n-Src loops, whose sequences vary among different SH3 domains (7-9).According to this description, the recognition of proline-rich sequences by SH3 domains, based primarily in the burial of hydrophobic surfaces in the ligand and SH3-binding site (7, 9), would be expected to present a thermodynamic signature dominated by the hydrophobic effect, with the main dr...

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Section: Discussionmentioning

confidence: 95%

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confidence: 99%

Role of Interfacial Water Molecules in Proline-rich Ligand Recognition by the Src Homology 3 Domain of Abl

Palencia

Cámara-Artigas

Pisabarro

et al. 2010

Journal of Biological Chemistry

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“…Nonsteroid ligands such as bicalutamide (44) or its S-1 agonist analog (43) extend ether linked parasubstituted phenyl groups into an open channel between the H-bonds between protein groups and buried water molecules can stabilize structure through compensatory changes in enthalpy and entropy (65) as might occur for the H-bonding projections of Gln-711, Arg-752, Met-745, His-874, and Tyr-874. Such water-mediated H-bonds can provide favorable enthalpy but less favorable entropy than direct H-bonds in protein-ligand interactions (64,66). Deeply buried structured water molecules engaged in multiple H-bonds increase protein flexibility and vibrational entropy, whereas direct protein-mediated H-bonds provide a more rigid structure with fewer degrees of freedom compared with water-mediated H-bonds (67).…”

Section: Discussionmentioning

confidence: 99%

Modulation of Androgen Receptor Activation Function 2 by Testosterone and Dihydrotestosterone

Askew

Gampe

Stanley

et al. 2007

Journal of Biological Chemistry

181

166

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The androgen receptor (AR) is transcriptionally activated by high affinity binding of testosterone (T) or its 5␣-reduced metabolite, dihydrotestosterone (DHT), a more potent androgen required for male reproductive tract development. The molecular basis for the weaker activity of T was investigated by determining T-bound ligand binding domain crystal structures of wild-type AR and a prostate cancer somatic mutant complexed with the AR FXXLF or coactivator LXXLL peptide. Nearly identical interactions of T and DHT in the AR ligand binding pocket correlate with similar rates of dissociation from an AR fragment containing the ligand binding domain. However, T induces weaker AR FXXLF and coactivator LXXLL motif interactions at activation function 2 (AF2). Less effective FXXLF motif binding to AF2 accounts for faster T dissociation from full-length AR. T can nevertheless acquire DHT-like activity through an AR helix-10 H874Y prostate cancer mutation. The Tyr-874 mutant side chain mediates a new hydrogen bonding scheme from exterior helix-10 to backbone protein core helix-4 residue Tyr-739 to rescue T-induced AR activity by improving AF2 binding of FXXLF and LXXLL motifs. Greater AR AF2 activity by improved core helix interactions is supported by the effects of melanoma antigen gene protein-11, an AR coregulator that binds the AR FXXLF motif and targets AF2 for activation. We conclude that T is a weaker androgen than DHT because of less favorable T-dependent AR FXXLF and coactivator LXXLL motif interactions at AF2. The androgen receptor (AR)2 is a member of the nuclear receptor superfamily of ligand-activated transcription factors. Androgen activation of AR regulates prostate growth, bone and muscle mass, and spermatogenesis and is a predisposing factor in prostate cancer. AR mediates transcriptional activity in response to two biologically active androgens that bind AR with similar high affinity (1). Testosterone (T) is the major circulating androgen secreted by the testis and the active androgen in muscle. Dihydrotestosterone (DHT), the 5␣-reduced metabolite of T, is a more potent androgen required for male reproductive tract development.AR has a modular structure composed of an NH 2 -terminal transactivation domain, central DNA binding domain, linker hinge region, and carboxyl-terminal ligand binding domain (LBD) (2). Like other nuclear receptor family members (3), AR has two major activation domains. Androgen-induced AR transcriptional activity depends on activation function 1 in the largely unstructured NH 2 -terminal region (2) and activation function 2 (AF2), a highly ordered hydrophobic surface in the LBD that requires androgen binding for its structural integrity (4).The degree to which AF2 contributes to overall AR activity depends on multiple competing factors. Unlike other nuclear receptors, AR AF2 binds a number of LXXLL-related motifs. Important among these is the AR NH 2 -terminal FXXLF motif 23 FQNLF 27 that binds AF2 in an androgen-dependent and -specific manner. AR FXXLF motif binding to AF2 is the bas...

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“…While the orientation of residue 266 depends on the type of side chain, the structure of this hydrogen bond network is conserved: upon replacing Thr266 with arginine, the side chain and the water molecule change places, while the positions of CWM3 and of residue 172 are maintained. The role of conserved waters in maintaining hydrogen bond networks has been demonstrated previously for protein-protein interfaces and protein-ligand and protein-cofactor binding (4,35,41). The hydrogen bond network of CWM3 is an example of an intraprotein network which is stable upon side chain substitution between protein families.…”

Section: Discussionmentioning

confidence: 70%

Conserved Water Molecules Stabilize the Ω-Loop in Class A β-Lactamases

Bös

Pleiss

2008

Antimicrob Agents Chemother

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A set of 49 high-resolution (<2.2 Å) structures of the TEM, SHV, and CTX-M class A ␤-lactamase families was systematically analyzed to investigate the role of conserved water molecules in the stabilization of the ⍀-loop. Overall, 13 water molecules were found to be conserved in at least 45 structures, including two water positions which were found to be conserved in all structures. Of the 13 conserved water molecules, 6 are located at the ⍀-loop, forming a dense cluster with hydrogen bonds to residues at the ⍀-loop as well as to the rest of the protein. This layer of conserved water molecules is packed between the ⍀-loop and the rest of the protein and acts as structural glue, which could reduce the flexibility of the ⍀-loop. A correlation between conserved water molecules and conserved protein residues could in general not be detected, with the exception of the conserved water molecules at the ⍀-loop. Furthermore, the evolutionary relationship between the three families, derived from the number of conserved water molecules, is similar to the relationship derived from phylogenetic analysis.␤-Lactamases (EC 3.5.2.6) hydrolyze the ␤-lactam ring and therefore are the main cause for resistance against ␤-lactam antibiotics. A sequence-based classification scheme was initially proposed by Ambler (1) and soon thereafter was extended by others (14, 28). Class A ␤-lactamases include the plasmid-borne TEM, SHV, and CTX-M ␤-lactamases, which are found most commonly in gram-negative bacteria. The TEM and SHV ␤-lactamase families are closely related (with a sequence identity of 67%), while the CTX-M family is more distant (with a sequence identity of 40% to TEM and SHV) (36). The members of the TEM and SHV families differ by only one to seven amino acids, while the members of the CTX-M family are more diverse (80% sequence identity) (13). Because of their importance in antibiotic resistance, the enzymes of class A ␤-lactamases are well studied and characterized (2,6,10,20,21,30), and a vast amount of crystallographic data is available. While the sequence similarity between the protein families is moderate, the structures of all TEM, SHV, and CTX-M ␤-lactamases show a remarkably high similarity. The enzymes are composed of two domains that are closely packed together: an all-␣ domain, consisting of eight ␣-helices, and an ␣/␤ domain, consisting of three ␣-helices and five ␤-sheets (15). The active site cavity is part of the interface between the two domains and is limited by the ⍀-loop. The ⍀-loop (residues 161 to 179) is a conserved structural element in all class A ␤-lactamases and is directly involved in the catalytic reaction of the enzymes because it positions the general base Glu166. Its conformation is anchored by a highly conserved salt bridge formed between Arg164 and Asp179 (19,37,38). However, the long ⍀-loop has only a few contacts with the rest of the protein and was therefore speculated to be a flexible element (20). Indeed, a molecular dynamics study (32) attributes some flexibility to a part of the ⍀-loop. How...

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Thermodynamic consequences of disrupting a water‐mediated hydrogen bond network in a protein:pheromone complex

Cited by 43 publications

References 42 publications

Role of Interfacial Water Molecules in Proline-rich Ligand Recognition by the Src Homology 3 Domain of Abl

Role of Interfacial Water Molecules in Proline-rich Ligand Recognition by the Src Homology 3 Domain of Abl

Modulation of Androgen Receptor Activation Function 2 by Testosterone and Dihydrotestosterone

Conserved Water Molecules Stabilize the Ω-Loop in Class A β-Lactamases

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