title: Spectral and kinetic characterization of 7,8-diaminopelargonic acid synthase from Mycobacterium tuberculosis.
abstract: The indispensability of biotin for crucial processes like lipid biosynthesis coupled to the absence of the biotin biosynthesis pathway in humans make the enzymes of this pathway, attractive targets for development of novel drugs against numerous pathogens including M. tuberculosis. We report the spectral and kinetic characterization of the Mycobacterium tuberculosis 7,8-Diaminopelargonic acid (DAPA) synthase, the second enzyme of the biotin biosynthesis pathway. In contrast to the E. coli enzyme, no quinonoid intermediate was detected during the steady state reaction between the enzyme and S-adenosyl-L-methionine (SAM). The second order rate constant for this half of the reaction was determined to be 1.75 +/- 0.11 M-1s-1. The Km values for 7-keto-8-aminopelargonic acid (KAPA) and SAM are 2.83 microM and 308.28 microM, respectively whereas the Vmax and kcat values for the enzyme are 0.02074 micromoles/min/ml and 0.003 s-1, respectively. Our initial studies pave the way for further detailed mechanistic and kinetic characterization of the enzyme.
authors.affiliation: Molecular Biophysics Unit, Indian Institute of Science, Bangalore, India.
authors.forename: Vikrant M
authors.initials: VM
authors.lastname: Bhor
authors.forename: Sagarika
authors.initials: S
authors.lastname: Dev
authors.forename: Ganga Ramu
authors.initials: GR
authors.lastname: Vasanthakumar
authors.forename: Avadhesha
authors.initials: A
authors.lastname: Surolia
mesh_terms: D002853:Chromatography, Liquid; D007700:Kinetics; D009169:Mycobacterium tuberculosis / Q000201:enzymology*; D011732:Pyridoxal Phosphate / Q000378:metabolism; D011733:Pyridoxamine / Q000031:analogs & derivatives / Q000378:metabolism; D012436:S-Adenosylmethionine / Q000378:metabolism; D021241:Spectrometry, Mass, Electrospray Ionization; D013057:Spectrum Analysis; D000637:Transaminases / Q000737:chemistry* / Q000378:metabolism*
chemical_list: D011732:Pyridoxal Phosphate; D011733:Pyridoxamine; D012436:S-Adenosylmethionine; C020029:7,8-diaminopelargonic acid aminotransferase; D000637:Transaminases; C010627:pyridoxamine phosphate