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@ARTICLE{Hwang:41869,
author = {Hwang, J.-Y. and Schlesinger, R. and Koch, K.-W.},
title = {{I}rregular dimerization of guanylate cyclase-activating
protein 1 mutants causes loss of target activation},
journal = {The FEBS journal},
volume = {271},
issn = {0014-2956},
address = {Oxford [u.a.]},
publisher = {Wiley-Blackwell},
reportid = {PreJuSER-41869},
pages = {3785 - 3793},
year = {2004},
note = {Record converted from VDB: 12.11.2012},
abstract = {Guanylate cyclase-activating proteins (GCAPs) are neuronal
calcium sensors that activate membrane bound guanylate
cyclases (EC 4.6.1.2.) of vertebrate photoreceptor cells
when cytoplasmic Ca2+ decreases during illumination. GCAPs
contain four EF-hand Ca2+-binding motifs, but the first
EF-hand is nonfunctional. It was concluded that for GCAP-2,
the loss of Ca2+-binding ability of EF-hand 1 resulted in a
region that is crucial for targeting guanylate cyclase
[Ermilov, A.N., Olshevskaya, E.V. $\&$ Dizhoor, A.M. (2001)
J. Biol. Chem.276, 48143-48148]. In this study we tested the
consequences of mutations in EF-hand 1 of GCAP-1 with
respect to Ca2+ binding, Ca2+-induced conformational changes
and target activation. When the nonfunctional first EF-hand
in GCAP-1 is replaced by a functional EF-hand the chimeric
mutant CaM-GCAP-1 bound four Ca2+ and showed similar
Ca2+-dependent changes in tryptophan fluorescence as the
wild-type. CaM-GCAP-1 neither activated nor interacted with
guanylate cyclase. Size exclusion chromatography revealed
that the mutant tended to form inactive dimers instead of
active monomers like the wild-type. Critical amino acids in
EF-hand 1 of GCAP-1 are cysteine at position 29 and proline
at position 30, as changing these to glycine was sufficient
to cause loss of target activation without a loss of
Ca2+-induced conformational changes. The latter mutation
also promoted dimerization of the protein. Our results show
that EF-hand 1 in wild-type GCAP-1 is critical for providing
the correct conformation for target activation.},
keywords = {Amino Acid Sequence / Animals / Calcium: metabolism /
Calcium-Binding Proteins: chemistry / Calcium-Binding
Proteins: genetics / Calcium-Binding Proteins: isolation
$\&$ purification / Calcium-Binding Proteins: metabolism /
Chromatography, Gel / Cysteine: metabolism / Dimerization /
EF Hand Motifs: genetics / Escherichia coli: genetics /
Guanylate Cyclase-Activating Proteins / Light Signal
Transduction / Molecular Sequence Data / Mutation / Proline:
metabolism / Protein Conformation / Reactive Oxygen Species:
metabolism / Recombinant Proteins: chemistry / Recombinant
Proteins: metabolism / Sequence Homology, Amino Acid /
Spectrometry, Fluorescence / Calcium-Binding Proteins (NLM
Chemicals) / Guanylate Cyclase-Activating Proteins (NLM
Chemicals) / Reactive Oxygen Species (NLM Chemicals) /
Recombinant Proteins (NLM Chemicals) / Proline (NLM
Chemicals) / Cysteine (NLM Chemicals) / Calcium (NLM
Chemicals) / J (WoSType)},
cin = {IBI-1 / IBI-2},
ddc = {540},
cid = {I:(DE-Juel1)VDB57 / I:(DE-Juel1)VDB58},
pnm = {Neurowissenschaften},
pid = {G:(DE-Juel1)FUEK255},
shelfmark = {Biochemistry $\&$ Molecular Biology},
typ = {PUB:(DE-HGF)16},
pubmed = {pmid:15355355},
UT = {WOS:000223711400017},
doi = {10.1111/j.1432-1033.2004.04320.x},
url = {https://juser.fz-juelich.de/record/41869},
}