CLONING OF GLUTARYL-COA DEHYDROGENASE CDNA, AND EXPRESSION OF WILD-TYPE AND MUTANT ENZYMES IN ESCHERICHIA-COLI

Citation
Si. Goodman et al., CLONING OF GLUTARYL-COA DEHYDROGENASE CDNA, AND EXPRESSION OF WILD-TYPE AND MUTANT ENZYMES IN ESCHERICHIA-COLI, Human molecular genetics, 4(9), 1995, pp. 1493-1498
Citations number
26
Language
INGLESE
art.tipo
Article
Categorie Soggetti
Genetics & Heredity",Biology
Journal title
ISSN journal
0964-6906
Volume
4
Issue
9
Year of publication
1995
Pages
1493 - 1498
Database
ISI
SICI code
0964-6906(1995)4:9<1493:COGDCA>2.0.ZU;2-7
Abstract
We have cloned, sequenced, and expressed cDNAs encoding wild type huma n glutaryl-CoA dehydrogenase subunit, and have expressed a mutant enzy me found in a patient with glutaric acidemia type I. The mutant protei n is expressed at the same level as the wild type in Escherichia coil, but has less than 1% of the activity of wild-type dehydrogenase. We a lso present evidence that the glutaryl-CoA dehydrogenase transcript is alternatively spliced in human fibroblasts and liver; the alternative ly spliced mRNA, when expressed in E.coli, encodes a stable but inacti ve protein. Purified expressed human glutaryl-CoA dehydrogenase has ki netic constants similar to those of the previously purified porcine de hydrogenase. The primary translation product from in vitro transcribed glutaryl-CoA dehydrogenase mRNA is translocated into mitochondria and processed in the same manner as most other nuclear-encoded mitochondr ial proteins, Human glutaryl-CoA dehydrogenase shows 53% sequence simi larity to porcine medium chain acyl-CoA dehydrogenase, and these simil arities were utilized to predict structure-function relationships in g lutaryl-CoA dehydrogenase.