Article,
A new Azotobacter vinelandii mannuronan C-5-epimerase gene (algG) is part of an alg gene cluster physically organized in a manner similar to that in Pseudomonas aeruginosa.
J. Bacteriol., 178 (20): 5884-9 (1996)
Abstract
Alginate is an unbranched polysaccharide composed of the two sugar
residues beta-D-mannuronic acid (M) and alpha-L-guluronic acid (G).
The M/G ratio and sequence distribution in alginates vary and are
of both biological and commercial significance. We have previously
shown that a family of highly related mannuronan C-5-epimerase genes
(algE1 to -E5) controls these parameters in Azotobacter vinelandii,
by catalyzing the Ca2+-dependent conversion of M to G at the polymer
level. In this report, we describe the cloning and expression of
a new A. vinelandii epimerase gene (here designated algG), localized
29 nucleotides downstream of the previously described gene algJ.
Sequence alignments show that algG does not belong to the same class
of genes as algE1 to -E5 but that it shares 66% sequence identity
with a previously described mannuronan C-5-epimerase gene (also designated
algG) from Pseudomonas aeruginosa. A. vinelandii algG was expressed
in Escherichia coli, and the enzyme was found to catalyze epimerization
in the absence of Ca2+, although the presence of the cation stimulated
the activity moderately. Surprisingly, all activity was blocked by
Zn2+. P. aeruginosa AlgG has been reported to contain an N-terminal
export signal sequence which is cleaved off during expression in
E. coli. This does not happen with A. vinelandii AlgG, which appears
to be produced at least partly in an insoluble form when expressed
at high levels in E. coli. DNA sequencing analyses of the regions
flanking algG suggest that the gene is localized in a cluster of
genes putatively involved in alginate biosynthesis, and the organization
of this cluster appears to be the same as previously described for
P. aeruginosa.
Tags
- *genes,
- *multigene
- ;
- acid
- aeruginosa/genetics
- alginates/metabolism
- amino
- analysis,
- azotobacter
- bacterial
- base
- calcium/pharmacology
- carbohydrate
- cations,
- cloning,
- coli/genetics
- comparative
- data
- divalent/pharmacology
- dna
- epimerases/*genetics/metabolism
- escherichia
- family
- gov't
- homology,
- molecular
- non-u.s.
- proteins/*genetics/metabolism
- proteins/metabolism
- pseudomonas
- recombinant
- research
- sequence
- species
- specificity
- study
- support,
- vinelandii/enzymology/*genetics
- zinc/pharmacology
