scholarly journals Purification and characterization of the acyltransferase involved in biosynthesis of the major mycobacterial cell envelope glycolipid – Monoacylated phosphatidylinositol dimannoside

2014 ◽  
Vol 100 ◽  
pp. 33-39 ◽  
Author(s):  
Zuzana Svetlíková ◽  
Peter Baráth ◽  
Mary Jackson ◽  
Jana Korduláková ◽  
Katarína Mikušová
Keyword(s):  
Cell Envelope ◽  
Purification And Characterization ◽  
Mycobacterial Cell

scholarly journals Purification and Characterization of the Bacterial UDP-GlcNAc:Undecaprenyl-Phosphate GlcNAc-1-Phosphate Transferase WecA

2008 ◽  
Vol 190 (21) ◽  
pp. 7141-7146 ◽  
Author(s):  
Bayan Al-Dabbagh ◽  
Dominique Mengin-Lecreulx ◽  
Ahmed Bouhss
Keyword(s):  
Cell Envelope ◽  
Functional Characterization ◽  
Integral Membrane Protein ◽  
Minimal Length ◽  
Purification And Characterization ◽  
Effects Of Ph ◽  
Undecaprenyl Phosphate ◽  
First Time ◽  
Lipid Substrate

ABSTRACT To date, the structural and functional characterization of proteins belonging to the polyprenyl-phosphate N-acetylhexosamine-1-phosphate transferase superfamily has been relentlessly held back by problems encountered with their overexpression and purification. In the present work and for the first time, the integral membrane protein WecA that catalyzes the transfer of the GlcNAc-1-phosphate moiety from UDP-GlcNAc onto the carrier lipid undecaprenyl phosphate, yielding undecaprenyl-pyrophosphoryl-GlcNAc, the lipid intermediate involved in the synthesis of various bacterial cell envelope components, was overproduced and purified to near homogeneity in milligram quantities. An enzymatic assay was developed, and the kinetic parameters of WecA as well as the effects of pH, salts, cations, detergents, and temperature on the enzyme activity were determined. A minimal length of 35 carbons was required for the lipid substrate, and tunicamycin was shown to inhibit the enzyme at submicromolar concentrations.

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