title: First ever isolation of bacterial prolipoprotein diacylglyceryl transferase in single step from Lactococcus lactis.
abstract: The unique bacterial enzyme phosphatidylglycerol: prolipoprotein diacylglyceryl transferase (Lgt) is the least studied enzyme of the ubiquitous bacterial lipoprotein synthetic pathway, mostly due to the low abundance of the enzyme. So far, Lgt has been studied to a limited extent in gram-negative bacteria, mainly in Escherichia coli. We, for the first time, report the isolation of an adequate amount of Lgt from the gram-positive lactic acid bacteria, Lactococcus lactis and compare this wild-type bacterial enzyme with the E. coli enzyme. The L. lactis Lgt, when purified by cationic-exchange chromatography, showed a 20-fold increase in the specific activity compared to that of the load, and 75% of the total Lgt activity loaded was recovered. Kinetically, L. lactis Lgt was found to be similar to the E. coli enzyme with matching K(m) and V(max), whereas the specific activity of the L. lactis enzyme was about 20 times less than that of the E. coli enzyme. Comparative bioinformatic analysis of L. lactis, E. coli and Staphylococcus aureus Lgt revealed that the conserved and catalytically important His-103 residue in E. coli Lgt, was altered to Tyr in L. lactis. Investigations showed that other bacteria where this alteration is visible, form a diversion within the gram-positive bacteria in evolution. Further analysis revealed Mycobacterium smegmatis to be the species which evolved with the alteration of His to Tyr.
authors.affiliation: Centre for Biotechnology, Anna University, Chennai 600 025, India.
authors.forename: Sanchari
authors.initials: S
authors.lastname: Banerjee
authors.forename: Krishnan
authors.initials: K
authors.lastname: Sankaran
mesh_terms: D000595:Amino Acid Sequence; D001426:Bacterial Proteins / Q000096:biosynthesis* / Q000737:chemistry / Q000235:genetics / Q000302:isolation & purification*; D002852:Chromatography, Ion Exchange; D019295:Computational Biology; D004926:Escherichia coli / Q000201:enzymology / Q000235:genetics / Q000378:metabolism; D005960:Glucosides / Q000737:chemistry / Q000378:metabolism; D007700:Kinetics; D013294:Lactococcus lactis / Q000201:enzymology* / Q000235:genetics / Q000378:metabolism; D008969:Molecular Sequence Data; D010802:Phylogeny; D016415:Sequence Alignment; D014166:Transferases / Q000096:biosynthesis* / Q000737:chemistry / Q000235:genetics / Q000302:isolation & purification*
chemical_list: D001426:Bacterial Proteins; D005960:Glucosides; C018619:octyl-beta-D-glucoside; D014166:Transferases; C042308:phosphatidylglycerol glyceryl transferase