Structural and Functional Characterization of the Histidine Phosphatase Domains of Human Sts-1 and Sts-2

Weijie Zhou, Yue Yin, Alexandra S. Weinheimer, Neena Kaur, Nick Carpino, Jarrod B. French

Research output: Contribution to journalArticlepeer-review

5 Scopus citations

Abstract

The suppressor of T cell signaling (Sts) proteins, Sts-1 and Sts-2, are homologous phosphatases that negatively regulate signaling pathways downstream of the T cell receptor. Functional inactivation of Sts-1 and Sts-2 in a murine model leads to resistance to systemic infection by the opportunistic pathogen, Candida albicans. This suggests that modulation of the host immune response by inhibiting Sts function may be a viable strategy for treating these deadly fungal pathogen infections. To better understand the molecular determinants of function and structure, we characterized the structure and steady-state kinetics of the histidine phosphatase domains of human Sts-1 (Sts-1HP) and Sts-2 (Sts-2HP). We determined the X-ray crystal structures of unliganded Sts-1HP and Sts-1HP in complex with sulfate to 2.5 and 1.9 Å, respectively, and the structure of Sts-2HP with sulfate to 2.4 Å. The steady-state kinetic analysis shows, as expected, that Sts-1HP has a phosphatase activity significantly higher than that of Sts-2HP and that the human and mouse proteins behave similarly. In addition, comparison of the phosphatase activity of full-length Sts-1 protein to Sts-1HP reveals similar kinetics, indicating that Sts-1HP is a functional surrogate for the native protein. We also tested known phosphatase inhibitors and determined that the SHP-1 inhibitor, PHPS1, is a potent inhibitor of Sts-1 (Ki = 1.05 ± 0.15 μM). Finally, we demonstrated that human Sts-1 has robust phosphatase activity against the substrate, Zap-70, in a cell-based assay. Collectively, these data suggest that the human Sts proteins are druggable targets and provide a structural basis for future drug development efforts.

Original languageEnglish (US)
Pages (from-to)4637-4645
Number of pages9
JournalBiochemistry
Volume56
Issue number35
DOIs
StatePublished - Sep 5 2017
Externally publishedYes

Bibliographical note

Funding Information:
*Phone: 631-632-8015. E-mail: jarrod.french@stonybrook.edu. *Phone: 631-632-4610. E-mail: nicholas.carpino@stonybrook. edu. ORCID Jarrod B. French: 0000-0002-6762-1309 Funding This work was supported by Stony Brook University, the National Heart, Lung, and Blood Institute of the National Institutes of Health under Grant U01HL127522 (J.B.F. and N.C.), and the Office of the Assistant Secretary of Defense for Health Affairs through the Peer Reviewed Medical Research Program under Grant W81XWH17-1-0147 (J.B.F. and N.C.). Additional support was provided by the Center for Biotechnology, a New York State Center for Advanced Technology, Stony Brook University, Cold Spring Harbor Laboratory, Brookhaven National Laboratory, and the Feinstein Institute for Medical Research. Notes The authors declare no competing financial interest.

Funding Information:
This work is based upon research conducted at the Northeastern Collaborative Access Team beamlines, which are funded by the National Institute of General Medical Sciences from the National Institutes of Health (P41 GM103403). The Pilatus 6M detector on the 24-ID-C beamline is funded by a NIH-ORIP HEI grant (S10 RR029205). This research used resources of the Advanced Photon Source, a U.S. Department of Energy (DOE) Office of Science User Facility operated for the DOE Office of Science by Argonne National Laboratory under Contract DE-AC02-06CH11357.

Publisher Copyright:
© 2017 American Chemical Society.

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