Identification and cDNA cloning of a novel human mosaic protein, LGN, based on interaction with G alpha i2.
We have used the yeast two-hybrid system to identify proteins that interact with the alpha-subunit of the heterotrimeric GTP-binding protein, Gi2. We screened a human B cell cDNA library with full-length G alpha i2 and isolated four positive colonies, one of which expressed the 44-kDa COOH terminus of a previously unrecognized 677-amino acid (aa) protein. A full-length clone was isolated from a HeLa cell cDNA library. The deduced protein contains 10 Leu-Gly-Asn repeats, and thus we named it LGN. Computer analysis indicates that LGN is a mosaic protein with seven repeated sequences of about 40 aa in length at its N-terminal end, and four repeated sequences of about 34 aa at its C-terminal end. Each of the two repeat regions shows substantial similarity to proteins found in other organisms. RT-PCR analysis of human tissues showed that the mRNA of LGN was ubiquitously expressed. The specificity of interaction between G alpha i2 and LGN was confirmed by an in vitro binding assay using recombinant proteins. These data indicate that the yeast two-hybrid system can identify novel proteins, such as LGN, that interact with G alpha proteins. As a mosaic protein, LGN shows similarity with portions of proteins from many species and thus may define a new protein family.
Mochizuki
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N
Department of Pharmacology, University of California, San Diego, La Jolla 92093-0636, USA.
Cho
G
G
Wen
B
B
Insel
P A
PA
D000595:Amino Acid Sequence; D000818:Animals; D001483:Base Sequence; D002352:Carrier Proteins / Q000235:genetics* / Q000378:metabolism; D003001:Cloning, Molecular; D003205:Computing Methodologies; D004247:DNA; D018076:DNA, Complementary; D019204:GTP-Binding Proteins / Q000378:metabolism*; D005982:Glutathione Transferase / Q000235:genetics; D006801:Humans; D047908:Intracellular Signaling Peptides and Proteins; D008969:Molecular Sequence Data; D011993:Recombinant Fusion Proteins / Q000235:genetics / Q000378:metabolism; D012441:Saccharomyces cerevisiae / Q000235:genetics; D017386:Sequence Homology, Amino Acid; D014018:Tissue Distribution
D002352:Carrier Proteins; D018076:DNA, Complementary; C102754:GPSM2 protein, human; D047908:Intracellular Signaling Peptides and Proteins; D011993:Recombinant Fusion Proteins; D004247:DNA; D005982:Glutathione Transferase; D019204:GTP-Binding Proteins