Regulation of UDP-3-O-[R-3-hydroxymyristoyl]-N-acetylglucosamine deacetylase in Escherichia coli. The second enzymatic step of lipid a biosynthesis.
The first enzyme of lipid A assembly in Escherichia coli is an acyltransferase that attaches an R-3-hydroxymyristoyl moiety to UDP-GlcNAc at the GlcNAc 3-OH. This reaction is reversible and thermodynamically unfavorable. The subsequent deacetylation of the product, UDP-3-O-[R-3-hydroxymyristoyl]-GlcNAc, is therefore the first committed step of lipid A biosynthesis. We now demonstrate that inhibition of either the acyltransferase or the deacetylase in living cells results in a 5-10-fold increase in the specific activity of the deacetylase in extracts prepared from such cells. Five other enzymes of the lipid A pathway are not affected. The elevated specific activity of deacetylase observed in extracts of lipid A-depleted cells is not accompanied by a significant change in the Km for the substrate, but is mainly an effect on Vmax. Western blots demonstrate that more deacetylase protein is indeed made. However, deacetylase messenger RNA levels are not significantly altered. Inhibition of lipid A biosynthesis must either stimulate the translation of available mRNA or slow the turnover of pre-existing deacetylase. In contrast, inhibition of 3-deoxy-D-manno-octulosonic acid (Kdo) biosynthesis has no effect on deacetylase specific activity. The underacylated lipid A-like disaccharide precursors that accumulate during inhibition of Kdo formation may be sufficient to exert normal feedback control.
Sorensen
P G
PG
Department of Biochemistry, Duke University Medical Center, Durham, North Carolina 27710, USA.
Lutkenhaus
J
J
Young
K
K
Eveland
S S
SS
Anderson
M S
MS
Raetz
C R
CR
D000581:Amidohydrolases / Q000378:metabolism*; D015152:Blotting, Northern; D003568:Cytidine Monophosphate / Q000031:analogs & derivatives / Q000096:biosynthesis; D004926:Escherichia coli / Q000201:enzymology*; D007700:Kinetics; D008050:Lipid A / Q000096:biosynthesis*; D008970:Molecular Weight; D010957:Plasmids / Q000378:metabolism; D012333:RNA, Messenger / Q000378:metabolism; D013379:Substrate Specificity; D013400:Sugar Acids
D008050:Lipid A; D012333:RNA, Messenger; D013400:Sugar Acids; C094610:cytidine-5'-monophosphate-3-deoxy-manno-octulosonic acid; D000581:Amidohydrolases; C097307:UDP-3-O-acyl-N-acetylglucosamine deacetylase; D003568:Cytidine Monophosphate