33 research outputs found
Cold-adapted RTX lipase from antarctic Pseudomonas sp. strain AMS8: isolation, molecular modeling and heterologous expression
A new strain of psychrophilic bacteria (designated strain AMS8) from Antarctic soil was screened for extracellular lipolytic activity and further analyzed using molecular approach. Analysis of 16S rDNA showed that strain AMS8 was similar to Pseudomonas sp. A lipase gene named lipAMS8 was successfully isolated from strain AMS8, cloned, sequenced and overexpressed in Escherichia coli. Sequence analysis revealed that lipAMS8 consist of 1,431 bp nucleotides that encoded a polypeptide consisting of 476 amino acids. It lacked an N-terminal signal peptide and contained a glycine- and aspartate-rich nonapeptide sequence at the C-terminus, which are known to be the characteristics of repeats-in-toxin bacterial lipases. Furthermore, the substrate binding site of lipAMS8 was identified as S207, D 255 and H313, based on homology modeling and multiple sequence alignment. Crude lipase exhibited maximum activity at 20 C and retained almost 50 % of its activity at 10 C. The molecular weight of lipAMS8 was estimated to be 50 kDa via sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE). The optimal expression level was attained using the recombinant plasmid pET32b/BL21(DE3) expressed at 15 C for 8 h, induced by 0.1 mM isopropyl β-D thiogalactoside (IPTG) at E. coli growth optimal density of 0.5
The Complete Genome Sequence of ‘Candidatus Liberibacter solanacearum’, the Bacterium Associated with Potato Zebra Chip Disease
Zebra Chip (ZC) is an emerging plant disease that causes aboveground decline of
potato shoots and generally results in unusable tubers. This disease has led to
multi-million dollar losses for growers in the central and western United States
over the past decade and impacts the livelihood of potato farmers in Mexico and
New Zealand. ZC is associated with ‘Candidatus
Liberibacter solanacearum’, a fastidious alpha-proteobacterium that is
transmitted by a phloem-feeding psyllid vector, Bactericera
cockerelli Sulc. Research on this disease has been hampered by a
lack of robust culture methods and paucity of genome sequence information for
‘Ca. L. solanacearum’. Here we present the
sequence of the 1.26 Mbp metagenome of ‘Ca. L.
solanacearum’, based on DNA isolated from potato psyllids. The coding
inventory of the ‘Ca. L. solanacearum’ genome was
analyzed and compared to related Rhizobiaceae to better
understand ‘Ca. L. solanacearum’ physiology and
identify potential targets to develop improved treatment strategies. This
analysis revealed a number of unique transporters and pathways, all potentially
contributing to ZC pathogenesis. Some of these factors may have been acquired
through horizontal gene transfer. Taxonomically, ‘Ca. L.
solanacearum’ is related to ‘Ca. L.
asiaticus’, a suspected causative agent of citrus huanglongbing, yet many
genome rearrangements and several gene gains/losses are evident when comparing
these two Liberibacter. species. Relative to ‘Ca. L.
asiaticus’, ‘Ca. L. solanacearum’ probably
has reduced capacity for nucleic acid modification, increased amino acid and
vitamin biosynthesis functionalities, and gained a high-affinity iron transport
system characteristic of several pathogenic microbes