Identification, cloning, and sequencing of a cDNA coding for rat gamma-glutamyl hydrolase.
Purified gamma-glutamyl hydrolase secreted from rat H35 hepatoma cells has been characterized as a diffuse band of 55 kDa on SDS-polyacrylamide gel electrophoresis that is converted to bands of 35 and 33 kDa after enzymatic removal of N-linked carbohydrate. Polyclonal antibodies against 55-kDa gamma-glutamyl hydrolase captured the enzyme activity and recognized the glycosylated and both deglycosylated forms of gamma-glutamyl hydrolase. A complete cDNA sequence of gamma-glutamyl hydrolase was obtained using degenerate oligonucleotides derived from peptide sequences, screening of a rat hepatoma cDNA library, and reverse transcription polymerase chain reaction. Based upon the deduced amino acid sequence the peptide component of gamma-glutamyl hydrolase had a molecular weight of 33,400. The results of amino acid analysis of the purified protein agreed with the deduced amino acid sequence in which there are seven potential asparagine-containing glycosylation sites.
Yao
R
R
Division of Molecular Medicine, Wadsworth Center, New York State Department of Health, Albany, New York 12201-0509, USA.
Nimec
Z
Z
Ryan
T J
TJ
Galivan
J
J
D000595:Amino Acid Sequence; D000818:Animals; D001483:Base Sequence; D003001:Cloning, Molecular; D018076:DNA, Complementary / Q000235:genetics; D008114:Liver Neoplasms, Experimental; D008969:Molecular Sequence Data; D010449:Peptide Mapping; D051381:Rats; D011623:gamma-Glutamyl Hydrolase / Q000235:genetics*
D018076:DNA, Complementary; D011623:gamma-Glutamyl Hydrolase