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Binding of guanylyl cyclase activating protein 1 (GCAP1) to retinal guanylyl cyclase (RetGC1) - The role of individual EF-hands
Journal article   Open access   Peer reviewed

Binding of guanylyl cyclase activating protein 1 (GCAP1) to retinal guanylyl cyclase (RetGC1) - The role of individual EF-hands

Igor V. Peshenko, Elena V. Olshevskaya and Alexander M. Dizhoor
The Journal of biological chemistry, v 283(31), pp 21747-21757
01 Aug 2008
DOI: https://doi.org/10.1074/jbc.M801899200
PMID: 18541533
Featured in Collection :   UN Sustainable Development Goals @ Drexel
url
https://doi.org/10.1074/jbc.m801899200View
Published, Version of Record (VoR)Open Access (License Unspecified),  Open
url
https://doi.org/10.1074/jbc.M801899200View
Published, Version of Record (VoR) Open

Abstract

Biochemistry & Molecular Biology Life Sciences & Biomedicine Science & Technology
Guanylyl cyclase activating protein 1 (GCAP1), after substitution of Ca2+ by Mg2+ in its EF-hands, stimulates photoreceptor guanylyl cyclase, RetGC1, in response to light. We inactivated metal binding in individual EF-hands of GCAP1 tagged with green fluorescent protein to assess their role in GCAP1 binding to RetGC1 in co-transfected HEK293 cells. When expressed alone, GCAP1 was uniformly distributed throughout the cytoplasm and the nuclei of the cells, but when co-expressed with either fluorescently tagged or non-tagged RetGC1, it co-localized with the cyclase in the membranes. The co-localization did not occur when the C-terminal portion of RetGC1, containing its regulatory and catalytic domains, was removed. Mutations that preserved Mg2+ binding in all three metal-binding EF-hands did not affect GCAP1 association with the cyclase in live cells. Locking EF-hand 4 in its apo-conformation, incapable of binding either Ca2+ or Mg2+, had no effect on GCAP1 association with the cyclase. In contrast to EF-hand 4, inactivation of EF-hand 3 reduced the efficiency of the co-localization, and inactivation of EF-hand 2 drastically suppressed GCAP1 binding to the cyclase. These results directly demonstrate that metal binding in EF-hand 2 is crucial for GCAP1 attachment to RetGC1, and that in EF-hand 3 it is less critical, although it enhances the efficiency of the GCAP1 docking on the target enzyme. Metal binding in EF-hand 4 has no role in the primary attachment of GCAP1 to the cyclase, and it only triggers the activator-to-inhibitor functional switch in GCAP1.

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Web of Science research areas
Biochemistry & Molecular Biology
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