Isolation and characterization of the aspartokinase and aspartate semialdehyde dehydrogenase operon from mycobacteria.
Diaminopimelic acid (DAP) is a major component of the peptidoglycan layer of the mycobacterial cell wall. The mycobacterial cell wall has been implicated as a potential virulence factor and is highly immunogenic. The pathway for biosynthesis of DAP may serve as a target in the design of antimycobacterial agents and construction of in vivo selection systems. Despite its significance, this biosynthetic pathway is poorly understood in mycobacteria. In order to develop a better understanding of mycobacterial DAP biosynthesis, the aspartate semialdehyde dehydrogenase (asd) genes of Mycobacterium smegmatis, bacille Calmette-Guerin (BCG), Mycobacterium avium, Mycobacterium leprae, and Mycobacterium tuberculosis were isolated. The M. smegmatis asd gene was isolated by complementation in Escherichia coli. This gene was then used to isolate the asd genes from other mycobacteria. The asd-complementing fragments from BCG and M. smegmatis were sequenced. An open reading frame upstream of the mycobacterial asd gene was identified as the mycobacterial aspartokinase gene (ask). Primer extension analysis revealed that the only transcriptional start in this region is found 5' of the ask gene. This observation indicates that the mycobacterial ask and asd genes are in an operon.
Cirillo
J D
JD
Howard Hughes Medical Institute, Department of Microbiology and Immunology, Albert Einstein College of Medicine, Bronx, New York 10461.
Weisbrod
T R
TR
Pascopella
L
L
Bloom
B R
BR
Jacobs
W R
WR
D000595:Amino Acid Sequence; D001222:Aspartate Kinase / Q000235:genetics*; D001223:Aspartate-Semialdehyde Dehydrogenase / Q000235:genetics*; D001426:Bacterial Proteins / Q000235:genetics*; D001483:Base Sequence; D016384:Consensus Sequence; D003960:Diaminopimelic Acid / Q000378:metabolism; D004926:Escherichia coli / Q000235:genetics; D005798:Genes, Bacterial; D005816:Genetic Complementation Test; D008969:Molecular Sequence Data; D009161:Mycobacterium / Q000201:enzymology / Q000235:genetics*; D016366:Open Reading Frames; D009876:Operon; D016415:Sequence Alignment; D017386:Sequence Homology, Amino Acid
D001426:Bacterial Proteins; D003960:Diaminopimelic Acid; D001223:Aspartate-Semialdehyde Dehydrogenase; D001222:Aspartate Kinase