Mapping the transition state of the WW domain β-sheet

Crane, Jason C.; Koepf, Edward K.; Gruebele, Martin

doi:10.1006/jmbi.2000.3665

Cited by 91 publications

(137 citation statements)

References 31 publications

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“…6). Because several mutagenesis studies (10,(43)(44)(45) have established the importance of the Trp 6 -Pro 32 interaction for the folding and the stability of WW domains, we think that a similar strong intermolecular Trp-Pro interaction is used to promote protein-protein interactions between the WW domain and its substrates (46). The molecular mechanisms of the Pin1 function very recently became clearer through an elegant study by Zhou et al (17), who showed that Pin1 enhances in vitro the dephosphorylation of Cdc25C by the major transPro-directed phosphatase PP2A.…”

Section: Discussionmentioning

confidence: 99%

1H NMR Study on the Binding of Pin1 Trp-Trp Domain with Phosphothreonine Peptides

Wintjens

Wieruszeski

Drobecq

et al. 2001

Journal of Biological Chemistry

122

143

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Trp-Trp (WW 1 ) domains are compact modules of 38 -40 amino acids long, folded into a three-stranded ␤ sheet, and found in single or tandem repeats in over 25 unrelated cellsignaling proteins (1, 2). Although their binding to prolinebased ligands is now well described (3, 4), little is known about their precise biological function. WW domains form a new family of protein-protein interaction modules targeted to proline residues, analogous to the Src homology (SH) 3 domains (5).Based on their proline-rich sequence binding specificities, WW domains are classified into five distinct groups (6). The N-terminal WW domain of the peptidyl-prolyl isomerase Pin1, an essential regulator at mitotic entry, is grouped among class IV domains, which bind peptides containing a proline residue preceded by a phosphoserine or a phosphothreonine (pSer/ pThr-Pro motif) (7). This latter motif is found in several mitotic Pin1-binding phosphoproteins (8), including the mitotic phosphatase Cdc25 (9, 10) and the microtubule-associated tau () protein (11).Site-directed mutagenesis experiments indicate that Pin1 binds phosphoproteins through its N-terminal WW domain and that the binding site mainly implicates the conserved residues Tyr 18 and Trp 29 (7, 11), which constitute a nearly flat hydrophobic area on the molecular surface of the WW domain. WW domains interacting with the core sequence PPXY, like Yesassociated protein, use the same hydrophobic surface for molecular recognition (3,4,12). However, this hydrophobic binding site alone is not likely to explain the phospho-dependent character of the ligand binding to the Pin1 WW domain.Recently the x-ray crystal structure of the full Pin1 protein bound to a doubly phosphorylated peptide (YpSPTpSPS) from the C-terminal domain (CTD) of RNA polymerase II was reported (13). The protein-peptide interactions are essentially limited to two regions on the WW domain surface. First, a phosphate binding pocket, encompassing the side chains of Ser 11 , Arg 12 , and the backbone amide of Arg 12 , anchors the interacting phosphate moiety via several hydrogen bonds. Second, the aromatic pair Tyr 18 -Trp 29 forms a molecular clamp that constrains the proline at position ϩ1 of the interacting phosphoserine. The peptide ligand binds to the Pin1 WW domain in a N-to C-terminal orientation, in contrast to the C-to N-terminal orientation that is found for two other proline-rich peptides in complex with a group I WW domain (3,14). Other proline recognition domains, such as the SH3 domain of the Caenorhabditis elegans signaling adaptor protein Sem5, can * This project was partly executed in the framework of the Génopole de Lille. The 600-MHz NMR facility used in this study was funded by the European community, the Région Nord-Pas de Calais, CNRS, and the Institut Pasteur de Lille. The costs of publication of this article were defrayed in part by the payment of page charges. This article must therefore be hereby marked "advertisement" in accordance with 18 U.S.C. Section 1734 solely to indicate this fact. 1 The ab...

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Section: Discussionmentioning

confidence: 99%

1H NMR Study on the Binding of Pin1 Trp-Trp Domain with Phosphothreonine Peptides

Wintjens

Wieruszeski

Drobecq

et al. 2001

Journal of Biological Chemistry

122

143

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“…WW domains have been used extensively in experimental and theoretical folding studies (1)(2)(3)(4)(12)(13)(14)(15)(16)(17)(18)(19). Traditional side-chain and amide-to-ester mutagenesis of Pin WW in conjunction with laser temperature-jump (T-jump) relaxation studies revealed that the folding rate was limited by nucleation at the loop 1 substructure (1)(2)(3)16), similar to the situation observed in SH3 domains (20,21).…”

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confidence: 97%

“…The selection criteria cannot always be optimized independently over the entire sequence of a protein. For the human Pin1 (hPin1) WW domain (Pin WW hereafter), we have shown that residues important for stability and folding rate are segregated in the sequence (1)(2)(3)(4). It is likely that functional selection criteria are predominant once minimal energetic criteria are met.…”

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confidence: 99%

Structure–function–folding relationship in a WW domain

Jäger

Zhang

Bieschke

et al. 2006

Proc. Natl. Acad. Sci. U.S.A.

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Protein folding barriers result from a combination of factors including unavoidable energetic frustration from nonnative interactions, natural variation and selection of the amino acid sequence for function, and͞or selection pressure against aggregation. The rate-limiting step for human Pin1 WW domain folding is the formation of the loop 1 substructure. The native conformation of this six-residue loop positions side chains that are important for mediating protein-protein interactions through the binding of Pro-rich sequences. Replacement of the wild-type loop 1 primary structure by shorter sequences with a high propensity to fold into a type-I ␤-turn conformation or the statistically preferred type-I G1 bulge conformation accelerates WW domain folding by almost an order of magnitude and increases thermodynamic stability. However, loop engineering to optimize folding energetics has a significant downside: it effectively eliminates WW domain function according to ligand-binding studies. The energetic contribution of loop 1 to ligand binding appears to have evolved at the expense of fast folding and additional protein stability. Thus, the two-state barrier exhibited by the wild-type human Pin1 WW domain principally results from functional requirements, rather than from physical constraints inherent to even the most efficient loop formation process.␤-turn ͉ ligand binding ͉ protein folding ͉ ␤-sheet ͉ protein function G lobular proteins evolve by mutation and selection. Selection criteria include function, and sufficient thermodynamic stability and folding rate to avoid sustained chaperone binding and proteasome degradation. The selection criteria cannot always be optimized independently over the entire sequence of a protein. For the human Pin1 (hPin1) WW domain (Pin WW hereafter), we have shown that residues important for stability and folding rate are segregated in the sequence (1-4). It is likely that functional selection criteria are predominant once minimal energetic criteria are met. Therefore, sequence evolution to enhance function may lead to a decrease in protein stability and folding rate compared with a sequence optimized for folding energetics.The hPin1 cell cycle regulatory proline (Pro) cis͞trans-isomerase is a two-domain protein (5). In its physiological role, the N-terminal WW domain binds Pro-rich ligands of the consensus sequence (pS͞pT)P, whereas the C-terminal domain catalyzes the Pro cis͞ trans-isomerization at the pS͞pT-P peptide bond. NMR solution studies show that the two domains, which are connected by a flexible solvated linker, interact only weakly before ligand binding (6, 7). The structure of the isolated Pin WW domain is virtually superimposable on that of the WW domain in the two-domain hPin1 protein (8). Moreover, Pin WW exhibits sufficient thermodynamic stability for biophysical analysis, folds rapidly, and retains its ligand-binding function (3, 9). These attributes, combined with sequence information on Ͼ150 WW domain family members (10, 11), makes Pin WW an excellent small model p...

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“…Later, the thermodynamics and kinetics of folding were compared between the following three WW domains (18): one from hYAP, one from murine forminbinding protein (FBP) 28, and a de novo-designed WW domain. All three of the aforementioned studies (16)(17)(18) reported singleexponential kinetics for folding, corresponding to an apparent two-state mechanism.…”

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confidence: 99%

“…Because of the attractiveness of WW domains as a model for ␤-sheet formation, they have been the focus of several previous folding studies. The initial study of these systems examined the thermodynamics and kinetics of folding of the human Yes-associated protein (hYAP) WW domain (16). A subsequent study made use of the more stable Pin WW domain, to characterize in detail the dependence of folding on temperature and denaturant (17).…”

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confidence: 99%

The structural basis for biphasic kinetics in the folding of the WW domain from a formin-binding protein: Lessons for protein design?

Karanicolas

Brooks

2003

Proc. Natl. Acad. Sci. U.S.A.

124

156

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The mechanism of formation of ␤-sheets is of great importance because of the significant role of such structures in the initiation and propagation of amyloid diseases. In this study we examine the folding of a series of three-stranded antiparallel ␤-sheets known as WW domains. Whereas other WW domains have been shown to fold with single-exponential kinetics, the WW domain from murine formin-binding protein 28 has recently been shown to fold with biphasic kinetics. By using a combination of kinetics and thermodynamics to characterize a simple model for this protein, the origins of the biphasic kinetics is found to lie in the fact that most of the protein is able to fold without requiring one of the ␤-hairpins to be correctly registered. The correct register of this hairpin is enforced by a surface-exposed hydrophobic contact, which is not present in other WW domains. This finding suggests the use of judiciously chosen surface-exposed hydrophobic pairs as a protein design strategy for enforcing the desired strand registry.␤-sheet ͉ ␤-strand ͉ negative design ͉ strand register A n understanding of the mechanism by which proteins reach their particular native state from the vast number of unfolded conformations will have broad-reaching implications, ranging from the prediction of protein structure from sequence to understanding diseases that originate from protein misfolding. Small model peptides and proteins have lent invaluable insight into the protein-folding process, as they afford simple systems in which the general features of folding may be elucidated (1, 2).Because of the local nature of the interactions, the formation of helices has proven considerably easier to understand using simple models (3-5) than has the formation of ␤-sheets. Accordingly, the principles that govern the formation of ␤-sheets are not well understood, despite the fact that the formation of intermolecular ␤-sheets is thought to be the crucial event in the initiation and propagation of amyloid diseases such as Alzheimer's disease (6) and spongiform encephalopathy (7).To further an understanding of the elements responsible for stability in ␤-sheets, de novo design methods have, in two cases, been used to construct three-stranded antiparallel ␤-sheets (8, 9), which have subsequently become the subject of both theoretical (10-12) and experimental (13, 14) folding studies. To ensure the generality of results toward natural proteins, however, we opt to study a series of three-stranded antiparallel ␤-sheet domains found in a variety of proteins: the WW domains (Fig. 1a).WW domains, which bind proline-rich peptide sequences, have been identified in Ͼ200 nonredundant proteins to date (15). Because of the attractiveness of WW domains as a model for ␤-sheet formation, they have been the focus of several previous folding studies. The initial study of these systems examined the thermodynamics and kinetics of folding of the human Yes-associated protein (hYAP) WW domain (16). A subsequent study made use of the more stable Pin WW domain, to characteriz...

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Mapping the transition state of the WW domain β-sheet

Cited by 91 publications

References 31 publications

1H NMR Study on the Binding of Pin1 Trp-Trp Domain with Phosphothreonine Peptides

1H NMR Study on the Binding of Pin1 Trp-Trp Domain with Phosphothreonine Peptides

Structure–function–folding relationship in a WW domain

The structural basis for biphasic kinetics in the folding of the WW domain from a formin-binding protein: Lessons for protein design?

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