Cloning, sequencing, and expression of Trypanosoma brucei dihydrolipoamide dehydrogenase.
A gene encoding dihydrolipoamide dehydrogenase was isolated from Trypanosoma brucei genomic DNA by using a combination of polymerase chain reaction and screening of a lambda EMBL3 library. The DNA sequence reveals that it encodes a protein of 478 amino acids (M(r) 49935) highly similar to previously sequenced dihydrolipoamide dehydrogenases. The gene was ligated into pMEX8 and expressed in an Escherichia coli mutant that lacks dihydrolipoamide dehydrogenase. Expression resulted in the appearance of dihydrolipoamide dehydrogenase activity concurrent with the production of a protein of the expected M(r) as determined by SDS/PAGE and Western blotting.
Else
A J
AJ
Department of Biochemistry, University of Bath, England.
Hough
D W
DW
Danson
M J
MJ
D000595:Amino Acid Sequence; D000818:Animals; D001483:Base Sequence; D003001:Cloning, Molecular; D003062:Codon; D004247:DNA / Q000737:chemistry; D008058:Dihydrolipoamide Dehydrogenase / Q000737:chemistry / Q000235:genetics*; D004926:Escherichia coli / Q000235:genetics; D015870:Gene Expression; D015723:Gene Library; D017125:Genes, Protozoan; D008969:Molecular Sequence Data; D008970:Molecular Weight; D010446:Peptide Fragments / Q000235:genetics; D014346:Trypanosoma brucei brucei / Q000201:enzymology*
D003062:Codon; D010446:Peptide Fragments; D004247:DNA; D008058:Dihydrolipoamide Dehydrogenase