Mutation of Lys‐75 affects calmodulin conformation

FEBS Letters

Gusev, Nikolai B.¹  Polyakova, Oxsana V.¹  Medvedeva, Marina V.¹  Watterson, D.Martin² 

[1] Department of Biochemistry, School of Biology, Moscow State University, Moscow 119899, Russia;Department of Molecular Pharmacology and Biological Chemistry, Northwestern University Medical School, Chicago, IL, USA

关键词: Calmodulin;    Mutant;    Limited proteolysis;    Mass-spectrometry;    CaM;    calmodulin;    EGTA;    ethyleneglycoltetraacetate;    MALDI-MS;    matrix assistant laser desorption mass-spectrometry;    SDS;    sodium dodecyl sulfate;   

DOI  :  10.1016/S0014-5793(99)00483-4

学科分类：生物化学/生物物理

来源: John Wiley & Sons Ltd.

【 摘 要 】

Some properties of synthetic calmodulin and its five mutants with replacement of Lys-75 were analyzed by means of electrophoresis, limited proteolysis and MALDI mass-spectrometry. A double mutant of calmodulin containing insert KGK between residues 80 and 81 and replacement of Lys-75 by Pro has a highly flexible central helix which is susceptible to trypsinolysis in the presence of Ca²⁺. Two mutants, K75P and K75E, having a distorted central helix demonstrate high resistance to trypsinolysis in the absence of Ca²⁺. Arg-90 and Arg-106 being the primary site of trypsinolysis of synthetic calmodulin are partially-protected in K75P and K75E mutants. The central helix of K75A and K75V mutants is stabilized by hydrophobic interactions between residues located in positions 71, 72 and 75. In the presence of Ca²⁺, the central helix of K75V is resistant to trypsinolysis. Mutations K75A and K75V decrease the rate of trypsinolysis of the central helix with a simultaneous increase of the rate of trypsinolysis in the C-terminal domain of calmodulin. It is concluded that the point mutation in the central helix has a long distance effect on the structure of calmodulin.

【 授权许可】

Unknown