Cloning, sequencing, and expression of Trypanosoma brucei dihydrolipoamide dehydrogenase
Open Access
- 3 March 1993
- journal article
- Published by Wiley in European Journal of Biochemistry
- Vol. 212 (2) , 423-429
- https://doi.org/10.1111/j.1432-1033.1993.tb17678.x
Abstract
A gene encoding dihydrolipoamide dehydrogenase was isolated from Trypanosoma brucei genomic DNA by using a combination of polymerase chain reaction and screening of a λ EMBL3 library. The DNA sequence reveals that it encodes a protein of 478 amino acids (Mr 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 Mr as determined by SDS/PAGE and Western blotting.Keywords
This publication has 32 references indexed in Scilit:
- Dihydrolipoamide dehydrogenase from Haloferax volcanii: gene cloning, complete primary structure, and comparison to other dihydrolipoamide dehydrogenasesBiochemistry and Cell Biology, 1992
- Cloning and sequence analysis of the genes encoding the dihydrolipoamide acetyltransferase and dihydrolipoamide dehydrogenase components of the pyruvate dehydrogenase multienzyme complex of Bacillus stearothermophilusEuropean Journal of Biochemistry, 1990
- Subcellular localisation of dihydrolipoamide dehydrogenase and detection of lipoic acid in bloodstream forms of Trypanosoma bruceiEuropean Journal of Biochemistry, 1990
- X-ray structure of lipoamide dehydrogenase from Azotobacter vinelandii determined by a combination of molecular and isomorphous replacement techniquesJournal of Molecular Biology, 1989
- Sequence analysis of the lpdV gene for lipoamide dehydrogenase of branched‐chain‐oxoacid dehydrogenase of Pseudomonas putidaEuropean Journal of Biochemistry, 1989
- Lipoamide dehydrogenase from Azotobacter vinelandiiEuropean Journal of Biochemistry, 1988
- Galactose- and maltose-stimulated lipoamide dehydrogenase activities related to the binding-protein-dependent transport of galactose and maltose in toluenized cells of Escherichia coliEuropean Journal of Biochemistry, 1986
- Genetic reconstruction and functional analysis of the repeating lipoyl domains in the pyruvate dehydrogenase multienzyme complex of Escherichia coliJournal of Molecular Biology, 1985
- The aerobic/anaerobic transition of glucose metabolism in Trypanosoma bruceiFEBS Letters, 1985
- A rapid and sensitive method for the quantitation of microgram quantities of protein utilizing the principle of protein-dye bindingAnalytical Biochemistry, 1976