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PhLPs and PhLOPs in the Phosducin Family of Gβγ Binding Proteins

Craft, Cheryl M. and Xu, Jun and Slepak, Vladlen Z. and Zhan-Poe, Xinyi and Zhu, Xuemei and Brown, Bruce and Lolley, Richard N. (1998) PhLPs and PhLOPs in the Phosducin Family of Gβγ Binding Proteins. Biochemistry, 37 (45). pp. 15758-15772. ISSN 0006-2960. https://resolver.caltech.edu/CaltechAUTHORS:20180213-083747762

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Abstract

In this study, we identify new isoforms of the retinal phosducin and investigate the expression of the phosducin family, showing that an isoform, PhLP1, has sequence homology with Phd and Gβγ binding capability, whereas two isoforms (phosducin-like orphan proteins, PhLOPs) share sequence homology with Phd but fail to bind Gβγ. Original identification of PhLP1 and the PhLOPs was from a human retina cDNA library, using a PCR product for library hybridization screening that contained a predicted functional epitope domain. The screen identified Phd and three related, but distinct, recombinants (PhLP1, PhLOP1, and PhLOP2). By RT-PCR, all isoforms are expressed in either retina or forskolin-stimulated Y79 retinoblastoma cells; however, the new isoforms are below the level of detection on Northern blot analysis. The predicted amino acid translation of each homologue revealed major differences, arising from either splice variants or gene duplication of Phd. To test the functional interaction of all phosducin isoforms with Gβγ in vitro, a glutathione S-transferase (GST) fusion protein was developed for each member. Biochemical interaction with purified retinal transducin Gβγ was verified for GST−Phd and demonstrated for GST−PhLP1; however, neither GST−PhLOP1 nor GST−PhLOP2 bound Gβγ. Comparable results were observed when the GST−phosducin fusion proteins selectively sequestered Gβγs from retinal extracts or when functional Gβγ interactions were assessed using surface plasmon resonance technology. Phosducin and its isoforms are widely distributed in body tissues where they may participate in signal transduction pathways. Phd and PhLP1 possess an 11-amino acid conserved epitope domain (TGPKGVINDWR) that controls the high-affinity binding of Gβγ; these isoforms are implicated in the G-protein signaling pathway. The phosducin-like orphan proteins (PhLOPs) fail to bind Gβγ, suggesting that the PhLOP isoforms may participate in still unidentified signaling pathways.


Item Type:Article
Related URLs:
URLURL TypeDescription
http://dx.doi.org/10.1021/bi980921aDOIArticle
https://pubs.acs.org/doi/full/10.1021/bi980921aPublisherArticle
Additional Information:© 1998 American Chemical Society. Received 23 April 1998. Published online 23 October 1998. Research support from the Mary D. Allen Endowment, NIH Grant NS28126 (C.M.C.), NIH Grant EY00395 (R.N.L.), the American Heart Association, Florida Affiliate (9603008) (V.Z.S.), Core Vision Research Center Grants EY03042 (Doheny Eye Institute) and AG12288 (M. I. Simon), the Michael P. Connell Foundation (C.M.C.), and the Neurogenetic Analysis Core (H.-J. Fülle) supported by the Howard Hughes Medical Institute Resources Grant (C.M.C.). C.M.C. is the Mary D. Allen Professor for Vision Research at the Doheny Eye Institute. This work is dedicated to Mary D. Allen for her generous support of vision research. We acknowledge Judge Murage and To Hoa Thai for excellent technical support.
Funders:
Funding AgencyGrant Number
NIHNS28126
NIHEY00395
American Heart Association9603008
NIHEY03042
NIHAG12288
Michael P. Connell FoundationUNSPECIFIED
Howard Hughes Medical Institute (HHMI)UNSPECIFIED
Doheny Eye InstituteUNSPECIFIED
Issue or Number:45
Record Number:CaltechAUTHORS:20180213-083747762
Persistent URL:https://resolver.caltech.edu/CaltechAUTHORS:20180213-083747762
Official Citation:PhLPs and PhLOPs in the Phosducin Family of Gβγ Binding Proteins Cheryl M. Craft, Jun Xu, Vladlen Z. Slepak, Xinyi Zhan-Poe, Xuemei Zhu, Bruce Brown, and Richard N. Lolley Biochemistry 1998 37 (45), 15758-15772 DOI: 10.1021/bi980921a
Usage Policy:No commercial reproduction, distribution, display or performance rights in this work are provided.
ID Code:84803
Collection:CaltechAUTHORS
Deposited By: Ruth Sustaita
Deposited On:13 Feb 2018 20:55
Last Modified:03 Oct 2019 19:22

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