Substrate Recognition Reduces Side-Chain Flexibility for Conserved Hydrophobic Residues in Human Pin1

Namanja, Andrew T.; Tao, Peng; Zintsmaster, John S.; Elson, Andrew; Shakour, Maria G.; Peng, Jeffrey W.

doi:10.1016/j.str.2007.01.014

Cited by 74 publications

(142 citation statements)

References 66 publications

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“…These alterations include both increases and decreases, with decreases (stiffening) dominating for the conduit residues that are less solvent accessible. This conduit overlaps with that observed in our previous study (6) of Pin1 interacting with a different substrate. The emergence of this conduit response for two substrates with different sequences and lengths (FFpSPR versus EQPLpTPVTDL) suggests that it is intrinsic to Pin1 substrate recognition.…”

Section: Discussionsupporting

confidence: 90%

“…4). These stiffening side chains formed an internal conduit, similar to that seen for Cdc25 (6). This was supported by rðFFpSPR;Cdc25Þ ¼ 0.66 AE 0.08.…”

Section: Ppiase Activity Is Modulated By Remote Interactions At the Dsupporting

confidence: 56%

“…The increased mobility of these side chains may reflect loosening at a "flexure point" to facilitate rearrangements of the catalytic loop upon ligand binding. Such rearrangements are evident in X-ray crystal structures (9,14), and implicated by our previous NMR exchange-broadening measurements (6).…”

Section: Ppiase Activity Is Modulated By Remote Interactions At the Dsupporting

confidence: 52%

“…To define more precisely the flexibility changes important for binding, we pinpointed the recurrent changes across four interactions including the three ligands herein, and our previous Cdc25 study (6). For example, ligand binding caused consistent loosening (ΔS 2 axis < 0) at methyl axes of A53β, V55γ 1 , T79γ 2 , and A85β.…”

Section: Ppiase Activity Is Modulated By Remote Interactions At the Dmentioning

confidence: 99%

“…Earlier structural studies of Pin1 revealed conformational changes upon substrate interaction, thus motivating flexibility-function studies of Pin1 (4-6). Our previous NMR deuterium relaxation studies of Pin1 mapped the changes in flexibility of methyl-bearing side chains caused by interaction with an established phospho-peptide (pT) substrate (6). The interaction caused both gains and losses in side-chain flexibility.…”

mentioning

confidence: 99%

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Stereospecific gating of functional motions in Pin1

Namanja

Wang

et al. 2011

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

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164

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Pin1 is a modular enzyme that accelerates the cis-trans isomerization of phosphorylated-Ser/Thr-Pro (pS/T-P) motifs found in numerous signaling proteins regulating cell growth and neuronal survival. We have used NMR to investigate the interaction of Pin1 with three related ligands that include a pS-P substrate peptide, and two pS-P substrate analogue inhibitors locked in the cis and trans conformations. Specifically, we compared the ligand binding modes and binding-induced changes in Pin1 side-chain flexibility. The cis and trans binding modes differ, and produce different mobility in Pin1. The cis-locked inhibitor and substrate produced a loss of side-chain flexibility along an internal conduit of conserved hydrophobic residues, connecting the domain interface with the isomerase active site. The trans-locked inhibitor produces a weaker conduit response. Thus, the conduit response is stereoselective. We further show interactions between the peptidyl-prolyl isomerase and Trp-Trp (WW) domains amplify the conduit response, and alter binding properties at the remote peptidyl-prolyl isomerase active site. These results suggest that specific input conformations can gate dynamic changes that support intraprotein communication. Such gating may help control the propagation of chemical signals by Pin1, and other modular signaling proteins.allostery | protein dynamics | ligand dynamics | protein evolution P hospho-serine/threonine-proline (pS/T-P) motifs are signaling motifs within intrinsically disordered loops of cell cycle proteins (1). The imide bond between the pS/T and P residues can adopt either the cis or trans conformation. These conformations differ in their susceptibility to kinases and phosphatases that propagate the chemical signals governing the cell cycle. Accordingly, the cell must regulate the cis/trans populations of these pS/T-P motifs to ensure proper signal routing.In this context, the peptidyl-prolyl isomerase Pin1 has emerged as a critical regulator (2, 3). Pin1 is a reversible enzyme that catalyzes the cis-trans isomerization of the pS/T-P imide linkages (2, 3) of other signaling proteins, such as CDC25C, p53, c-Myc, NF-kB, cyclin D1, and tau (3). Pin1 engages when external events, such as S/T (de)-phosphorylation, change the cis-trans equilibrium. Pin1 then catalyzes the cis-trans isomerization, thereby accelerating the approach to the new equilibrium (1).Pin1 is a modular protein of 163 residues consisting of a WW domain (1-39) and a larger peptidyl-prolyl isomerase (PPIase) domain (50-163) (Fig. 1). A flexible linker connects the two domains. Both domains are specific for pS/T-P motifs (1). The WW domain serves as a docking module, whereas catalysis is the sole province of the PPIase domain. Earlier structural studies of Pin1 revealed conformational changes upon substrate interaction, thus motivating flexibility-function studies of Pin1 (4-6). Our previous NMR deuterium relaxation studies of Pin1 mapped the changes in flexibility of methyl-bearing side chains caused by interaction with an establi...

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Section: Discussionsupporting

confidence: 90%

“…4). These stiffening side chains formed an internal conduit, similar to that seen for Cdc25 (6). This was supported by rðFFpSPR;Cdc25Þ ¼ 0.66 AE 0.08.…”

Section: Ppiase Activity Is Modulated By Remote Interactions At the Dsupporting

confidence: 56%

Section: Ppiase Activity Is Modulated By Remote Interactions At the Dsupporting

confidence: 52%

Section: Ppiase Activity Is Modulated By Remote Interactions At the Dmentioning

confidence: 99%

mentioning

confidence: 99%

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Stereospecific gating of functional motions in Pin1

Namanja

Wang

et al. 2011

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

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164

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Dynamics in Enzymes: Solution NMR Studies

Loria

2012

Encyclopedia of Magnetic Resonance

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Exploring the role of structure and dynamics in the function of chymotrypsin inhibitor 2

Whitley¹,

Lee²

2010

Proteins

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Increasing awareness of the possible role of internal dynamics in protein function has led to the development of new methods for experimentally characterizing protein dynamics across multiple time scales, especially using NMR spectroscopy. A few analyses of the conformational dynamics of proteins ranging from nonallosteric single domains to multi-domain allosteric enzymes are now available; however, demonstrating a connection between dynamics and function remains difficult on account of the comparative lack of studies examining both changes in dynamics and changes in function in response to the same perturbations. In previous work, we characterized changes in structure and dynamics on the ps-ns time scale resulting from hydrophobic core mutations in chymotrypsin inhibitor 2 and found that there are moderate, persistent global changes in dynamics in the absence of gross structural changes [Whitley MJ, et al. Biochemistry 2008; 47:8566-8576]. Here, we assay those and additional mutants for inhibitory ability toward the serine proteases elastase and chymotrypsin to determine the effects of mutation on function. Results indicate that core mutation has only a subtle effect on CI2 function. Using chemical shifts, we also studied the effect of complex formation on CI2 structure and found that perturbations are greatest at the complex interface but also propagate toward CI2’s hydrophobic core. The structure-dynamics-function data set completed here suggests that dynamics plays a limited role in the function of this small model system, although we do observe a correlation between nanosecond-scale reactive loop motions and inhibitory ability for mutations at one key position in the hydrophobic core.

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Substrate Recognition Reduces Side-Chain Flexibility for Conserved Hydrophobic Residues in Human Pin1

Cited by 74 publications

References 66 publications

Stereospecific gating of functional motions in Pin1

Stereospecific gating of functional motions in Pin1

Dynamics in Enzymes: Solution NMR Studies

Exploring the role of structure and dynamics in the function of chymotrypsin inhibitor 2

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