The Structural Basis for the Selectivity of Benzotriazole Inhibitors of PTP1B

Scapin, Giovanna; Patel, Sangita; Becker, Joseph W.; Wang, Qingping; Desponts, Caroline; Waddleton, Deena; Skorey, Kathryn; Cromlish, Wanda; Bayly, Christopher I.; Thérien, Michel; Gauthier, Jacques Yves; Li, Chun Sing; Lau, Cheuk K.; Ramachandran, Chidambaram; Kennedy, Brian P.; Asante‐Appiah, Ernest

doi:10.1021/bi035098j

Cited by 100 publications

(80 citation statements)

References 32 publications

(42 reference statements)

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“…Similarly, efforts to identify differences between TCPTP and PTP-1B by enzyme kinetic analysis did not reveal any discernible differences between the two enzymes that could be exploited for inhibitor design to achieve selectivity (23). Furthermore, a structural comparison of the three-dimensional crystal structures of both enzymes did not reveal obvious determinants of selectivity within the active site region.…”

Section: Resultsmentioning

confidence: 98%

“…The activity of all enzymes was assayed with fluorescein diphosphate (FDP) as substrate as previously reported (23,26). Kinetic constants were obtained by fitting the observed rates of reactions to the Michaelis-Menten equation with the aid of the non-linear curve fitting software program, Grafit 4.0.10 (Erithacus Software Inc.).…”

mentioning

confidence: 99%

“…Crystal Structure Determination-Crystals of the isolated catalytic domains of WT PTP-1B and L119V mutant in complex with compound 1 were obtained as previously reported (23). The data sets were collected from a single crystal using synchrotron radiation at the beamline 17ID in the facilities of the Industrial Macromolecular Crystallographic Association Collaborative Access Team (IMCA-CAT) at the Advanced Photon Source (Argonne National Laboratory, Argonne, IL).…”

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confidence: 99%

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Conformation-assisted Inhibition of Protein-tyrosine Phosphatase-1B Elicits Inhibitor Selectivity over T-cell Protein-tyrosine Phosphatase

Asante‐Appiah¹,

Patel

Desponts³

et al. 2006

Journal of Biological Chemistry

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PTP-1B represents an attractive target for the treatment of type 2 diabetes and obesity. Given the role that protein phosphatases play in the regulation of many biologically relevant processes, inhibitors against PTP-1B must be not only potent, but also selective. It has been extremely difficult to synthesize inhibitors that are selective over the highly homologous TCPTP. We have successfully exploited the conservative Leu 119 to Val substitution between the two enzymes to synthesize a PTP-1B inhibitor that is an order of magnitude more selective over TCPTP. Structural analyses of PTP-1B/inhibitor complexes show a conformation-assisted inhibition mechanism as the basis for selectivity. Such an inhibitory mechanism may be applicable to other homologous enzymes.The quest for agents that can intervene in type 2 diabetes continues to be a major research focus and challenge in many laboratories. As impaired insulin action is an underlying mechanism in type 2 diabetes, the insulin signaling pathway has naturally been the focus of research in attempts to identify suitable therapeutic target(s) for drug intervention against the disease. Insulin signaling begins with the activation of the insulin receptor (IR) via tyrosine phosphorylation and culminates in the uptake of glucose into cells by the glucose transporter, Glut4 (1). The activated IR must then be deactivated and returned to a basal state, a process that is believed to involve protein-tyrosine phosphatase-1B (PTP-1B).4 PTP-1B has been shown to directly interact with the activated insulin receptor (2-6). Disruption of the gene that codes for PTP-1B in mice results in sensitivity to insulin and also increased resistance to diet-induced obesity (7,8). This phosphatase is therefore a very attractive therapeutic target for the treatment not only of type 2 diabetes but also of obesity. Predictably, there has been an intense research effort by many groups to identify potent and selective inhibitors of PTP-1B (for recent reviews, see Refs. 9 and 10). PTP-1B belongs to the proteintyrosine phosphatase (PTP) superfamily of enzymes, which includes ϳ100 members involved in signal transduction and regulation of cellular processes such as growth, differentiation, and proliferation. Inhibitors against the enzyme must thus be not only potent but also selective. Sequence alignment of amino acids residues within the PTPs catalytic domain shows that, in general, all phosphatases have less than 40% identity to PTP-1B: a remarkable exception is the T-cell protein-tyrosine phosphatase (TCPTP), which has a 72% identity (11)(12)(13)(14). Inhibitors that target PTP-1B and incorporate previously identified selectivity determinants (15-17) are generally selective (Ͼ30-fold) over all PTPs tested but are equipotent on TCPTP. Although it is not evident that co-inhibition of TCPTP (with PTP-1B) will result in serious adverse effects, mice lacking the TCPTP gene die within 3-5 weeks after birth from defects in hematopoiesis and immune function (18). Hence, it is highly desirable to design PTP-1...

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Section: Resultsmentioning

confidence: 98%

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confidence: 99%

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confidence: 99%

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Conformation-assisted Inhibition of Protein-tyrosine Phosphatase-1B Elicits Inhibitor Selectivity over T-cell Protein-tyrosine Phosphatase

Asante‐Appiah¹,

Patel

Desponts³

et al. 2006

Journal of Biological Chemistry

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“…Similarly, we have reported on the crystal structure of difluorobenzotriazole inhibitors complexed with PTP1B and found that the two fluorine atoms are within van der Waals distance of Phe 182 and that they are hydrogen-bonded to a water molecule that also interacts with the phosphonate, the side chain nitrogen of Gln 266 , and the main chain nitrogen of Phe 182 (25). Examination of the crystal structures of PTP1B revealed that helix ␣7 interacts with the rest of the enzyme through an extensive series of hydrogen bonds with helix ␣3 and loop ␤9 -␤10 ( Fig.…”

Section: Tablementioning

confidence: 55%

“…1). The DFMP group of Inhibitor 1 engages the phosphatase by extensive hydrogen bonding of the phosphonate with the PTP loop (24,25). The two fluorine atoms are within van der Waals distance of the phenyl side chain of Phe 182 and are hydrogen-bonded through a water molecule to the side chain nitrogen of Gln 266 and the main chain nitrogen of Phe 182 (26,27).…”

Section: Use Of Yeast To Identify Amino Acid Determinants Of Ptp1b Inmentioning

confidence: 99%

Residues Distant from the Active Site Influence Protein-tyrosine Phosphatase 1B Inhibitor Binding

Montalibet

Skorey²,

McKay

et al. 2006

Journal of Biological Chemistry

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Regions of protein-tyrosine phosphatase (PTP) 1B that are distant from the active site yet affect inhibitor binding were identified by a novel library screen. This screen was based on the observation that expression of v-Src in yeast leads to lethality, which can be rescued by the coexpression of PTP1B. However, this rescue is lost when yeast are grown in the presence of PTP1B inhibitors. To identify regions of PTP1B (amino acids 1-400, catalytic domain plus 80-amino acid C-terminal tail) that can affect the binding of the difluoromethyl phosphonate (DFMP) inhibitor 7-bromo-6-difluoromethylphosphonate 3-naphthalenenitrile, a library coexpressing PTP1B mutants and v-Src was generated, and the ability of yeast to grow in the presence of the inhibitor was evaluated. PTP1B inhibitor-resistant mutations were found to concentrate on helix ␣7 and its surrounding region, but not in the active site. No resistant amino acid substitutions were found to occur in the C-terminal tail, suggesting that this region has little effect on active-site inhibitor binding. An in-depth characterization of a resistant substitution localizing to region ␣7 (S295F) revealed that this change minimally affected enzyme catalytic activity, but significantly reduced the potency of a panel of structurally diverse DFMP PTP1B inhibitors. This loss of inhibitor potency was found to be due to the difluoro moiety of these inhibitors because only the difluoro inhibitors were shifted. For example, the inhibitor potency of a monofluorinated or non-fluorinated analog of one of these DFMP inhibitors was only minimally affected. Using this type of library screen, which can scan the nearly full-length PTP1B sequence (catalytic domain and C-terminal tail) for effects on inhibitor binding, we have been able to identify novel regions of PTP1B that specifically affect the binding of DFMP inhibitors.A traditional approach to understanding the structural interactions between an inhibitor and enzyme is through crystallographic studies of the inhibitor-enzyme complex. Solving the three-dimensional structure of such complexes can identify the key residues that interact with the inhibitor, providing an understanding of the mechanism of inhibition. This is normally accompanied by site-directed mutagenesis of the interacting residues to validate their role in inhibitor binding and to determine the degree to which this interaction contributes to inhibitor potency. Such structures are also important in drug design, as they allow the identification of potential interactions that could improve inhibitor potency using further chemical modifications.Protein-tyrosine phosphatase (PTP) 3 1B (EC 3.1.3.48), a potential drug target for the treatment of diabetes and obesity (1, 2), has had a number of published inhibitor-enzyme complexes, and the determinants required for the binding of various inhibitors have been identified. Because these inhibitors all target the active site, the determinants identified are located in the active-site region. Fig. 4).Because of the static nature...

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Cu(I)/KOH‐Promoted Condensation between o‐Arylenediamines and Nitroarenes to Access 2‐Aryl‐2H‐Benzotriazoles

Gao

Huang

et al. 2020

Adv Synth Catal

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The Structural Basis for the Selectivity of Benzotriazole Inhibitors of PTP1B

Cited by 100 publications

References 32 publications

Conformation-assisted Inhibition of Protein-tyrosine Phosphatase-1B Elicits Inhibitor Selectivity over T-cell Protein-tyrosine Phosphatase

Conformation-assisted Inhibition of Protein-tyrosine Phosphatase-1B Elicits Inhibitor Selectivity over T-cell Protein-tyrosine Phosphatase

Residues Distant from the Active Site Influence Protein-tyrosine Phosphatase 1B Inhibitor Binding

Cu(I)/KOH‐Promoted Condensation between o‐Arylenediamines and Nitroarenes to Access 2‐Aryl‐2H‐Benzotriazoles

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