Bl. Hanzelka et al., MUTATIONAL ANALYSIS OF THE VIBRIO-FISCHERI LUXI POLYPEPTIDE - CRITICAL REGIONS OF AN AUTOINDUCER SYNTHASE, Journal of bacteriology, 179(15), 1997, pp. 4882-4887
Synthesis of the Vibrio fischeri autoinducer, a signal involved in the
cell density-dependent activation of bioluminescence, is directed by
the luxI gene product, The LuxI protein catalyzes the synthesis of N-a
cyl-homoserine lactones from S-adenosylmethionine and acylated-acyl ca
rrier protein, We have gained an appreciation of the LuxI regions and
amino acid residues involved in autoinducer synthesis by isolating and
analyzing mutations generated by random and site-specific mutagenesis
of luxI. By random mutagenesis we isolated 13 different single amino
acid substitutions in the LuxI polypeptide. Eleven of these substituti
ons resulted in no detectable autoinducer synthase activity, while the
remaining two amino acid substitutions resulted in reduced but detect
able activity, The substitutions that resulted in no detectable autoin
ducer synthase activity mapped to two small regions of LuxI. In Escher
ichia coli, wild-type luxI showed dominance over all of the mutations,
Because autoinducer synthesis has been proposed to involve formation
of a covalent bond between an acyl group and an active-site cysteine,
we constructed site-directed mutations that altered each of the three
cysteine residues in LuxI, All of the cysteine mutants retained substa
ntial activity as an autoinducer synthase in E, coli, Based on the ana
lysis of random mutations we propose a model in which there are two cr
itical regions of LuxI, at least one of which is an intimate part of a
n active site, and based on the analysis of site-directed mutations we
conclude that an active-site cysteine is not essential for autoinduce
r synthase activity.