Premium
The importance of Loop 7 for the activity of calcineurin
Author(s) -
Liu Ping,
Huang Chao,
Wang Hai-long,
Zhou Ke,
Xiao Fang-xiang,
Qun Wei
Publication year - 2004
Publication title -
febs letters
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.593
H-Index - 257
eISSN - 1873-3468
pISSN - 0014-5793
DOI - 10.1016/j.febslet.2004.09.082
Subject(s) - protein subunit , enzyme , calcineurin , mutant , active site , chemistry , loop (graph theory) , substrate (aquarium) , stereochemistry , enzyme activator , enzyme assay , mutation , biochemistry , biology , medicine , ecology , surgery , mathematics , combinatorics , transplantation , gene
Calcineurin (CN) is a heterodimer composed of a catalytic subunit (CNA) and a regulatory subunit (CNB). Loop 7 lies within the CNA catalytic domain. To investigate the role of Loop 7 in enzyme activity, we systematically examined all its residues by site‐directed deletion mutation. Our results show that the Loop 7 residues are important for enzyme activity. Besides deleting residues V314, Y315 or N316, enzyme activity also increased dramatically when residues D313 or K318 were deleted. In contrast, almost all activity was lost when L312 or N317 were deleted. Ni 2+ and Mn 2+ were effective activators for all active mutants. However, whereas the wild‐type enzyme was more efficiently activated by Ni 2+ than by Mn 2+ with 32 P‐labeled R II peptide as substrate, the reverse was true in all the mutants. We also found that the effect of Loop 7 on enzyme activity was substrate dependent, and involved interactions between Loop 7 residues and the unresolved part of the CN crystal structure near the auto‐inhibitory domain and catalytic site.