Comparative mechanistic and substrate specificity study of inositol polyphosphate 5-phosphatase Schizosaccharomyces pombe synaptojanin and SHIP2

Yuling Chi, Bo Zhou, Wei Qing Wang, Sung Kee Chung, Yong Uk Kwon, Young Hoon Ahn, Young Tae Chang, Yosuke Tsujishita, James H. Hurley, Zhong Yin Zhang

Research output: Contribution to journalArticle

36 Scopus citations

Abstract

Inositol-5-phosphatases are important enzymes involved in the regulation of diverse cellular processes from synaptic vesicle recycling to insulin signaling. We describe a comparative study of two representative inositol-5-phosphatases, Schizosaccharomyces pombe synaptojanin (SPsynaptojanin) and human SH2 domain-containing inositol-5-phosphatase SHIP2. We show that in addition to Mg2+, transition metals such as Mn2+, Co 2+, and Ni2+ are also effective activators of SPsynaptojanin. In contrast, Ca2+ and Cu2+ are inhibitory. We provide evidence that Mg2+ binds the same site occupied by Ca2+ observed in the crystal structure of SPsynaptojanin complexed with inositol 1,4-bisphosphate (Ins(1,4)P2). Ionizations important for substrate binding and catalysis are defined for the SPsynaptojanin-catalyzed Ins(1,4,5)P3 reaction. Kinetic analysis with four phosphatidylinositol lipids bearing a 5-phosphate and 54 water-soluble inositol phosphates reveals that SP-synaptojanin and SHIP2 possess much broader substrate specificity than previously appreciated. The rank order for SPsynaptojanin is Ins(2,4,5)P3 > phosphatidylinositol-4,5-bisphosphate (PtdIns(4,5)P2) ≈ Ins(4,5)P2 ≈ Ins(1,4,5)P 3 ≈ Ins(4,5,6)P3 > PtdIns(3,5)P2 ≈ PtdIns(3,4,5)P3 ≈ Ins(1,2,4,5)P4 ≈ Ins(1,3,4,5)P4 ≈ Ins-(2,4,5,6)P4 ≈ Ins(1,2,4,5,6)P5. The rank order for SHIP2 is Ins(1,2,3,4,5)P 5 > Ins(1,3,4,5)P4 > PtdIns(3,4,5)P4 ≈ PtdIns(3,5)P2 ≈ Ins(1,4,5,6)P4 ≈ Ins(2,4,5,6)P 4. Because inositol phosphate isomers elicit different biological activities, the extended substrate specificity for SPsynaptojanin and SHIP2 suggest that these enzymes likely have multiple roles in cell signaling and may regulate distinct pathways. The unique substrate specificity profiles and the importance of 2-position phosphate in binding also have important implications for the design of potent and selective SPsynaptojanin and SHIP2 inhibitors for pharmacological investigation.

Original languageEnglish (US)
Pages (from-to)44987-44995
Number of pages9
JournalJournal of Biological Chemistry
Volume279
Issue number43
DOIs
Publication statusPublished - Oct 22 2004

    Fingerprint

ASJC Scopus subject areas

  • Biochemistry
  • Molecular Biology
  • Cell Biology

Cite this