Residue 182 influences the second step of protein-tyrosine phosphatase-mediated catalysis

Ana K. Pedersen, Xiao Ling Guo, Karin B. Møller, Günther H. Peters, Henrik S. Andersen, Jette S. Kastrup, Steen B. Mortensen, Lars F. Iversen, Zhong Yin Zhang, Niels Peter H. Møller

Research output: Contribution to journalArticle

27 Scopus citations

Abstract

Previous enzyme kinetic and structural studies have revealed a critical role for Asp181 (PTP1B numbering) in PTP (protein-tyrosine phosphatase)-mediated catalysis. In the E-P (phosphoenzyme) formation step, Asp181 functions as a general acid, while in the E-P hydrolysis step it acts as a general base. Most of our understanding of the role of Asp 181 is derived from studies with the Yersinia PTP and the mammalian PTP1B, and to some extent also TC (T-cell)-PTP and the related PTPα and PTPε. The neighbouring residue 182 is a phenylalanine in these four mammalian enzymes and a glutamine in Yersinia PTP. Surprisingly, little attention has been paid to the fact that this residue is a histidine in most other mammalian PTPs. Using a reciprocal single-point mutational approach with introduction of His182 in PTP1B and Phe182 in PTPH1, we demonstrate here that His182-PTPs, in comparison with Phe 182-PTPs, have significantly decreased kcat values, and to a lesser degree, decreased kcat/Km values. Combined enzyme kinetic, X-ray crystallographic and molecular dynamics studies indicate that the effect of His182 is due to interactions with Asp 181 and with Gln262. We conclude that residue 182 can modulate the functionality of both Asp181 and Gln262 and therefore affect the E-P hydrolysis step of PTP-mediated catalysis.

Original languageEnglish (US)
Pages (from-to)421-433
Number of pages13
JournalBiochemical Journal
Volume378
Issue number2
DOIs
StatePublished - Mar 1 2004

Keywords

  • Enzyme kinetics
  • Molecular dynamics
  • PTP1B
  • PTPH1
  • Protein-tyrosine phosphatase
  • X-ray crystallography

ASJC Scopus subject areas

  • Biochemistry
  • Molecular Biology
  • Cell Biology

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    Pedersen, A. K., Guo, X. L., Møller, K. B., Peters, G. H., Andersen, H. S., Kastrup, J. S., Mortensen, S. B., Iversen, L. F., Zhang, Z. Y., & Møller, N. P. H. (2004). Residue 182 influences the second step of protein-tyrosine phosphatase-mediated catalysis. Biochemical Journal, 378(2), 421-433. https://doi.org/10.1042/BJ20030565