Document Detail


Functional analysis of the sialyltransferase complexes in Escherichia coli K1 and K92.
MedLine Citation:
PMID:  1735705     Owner:  NLM     Status:  MEDLINE    
Abstract/OtherAbstract:
The polysialyltransferase (polyST) structural gene, neuS, for poly alpha 2,8sialic acid (PSA) capsule synthesis in Escherichia coli K1 was previously mapped near the kps region 1 and 2 junction (S. M. Steenbergen and E. R. Vimr, Mol. Microbiol. 4:603-611, 1990). Present Southern and colony blot hybridization results confirmed that neuS was a region 2 locus and indicated apparent homology with neuS from E. coli K92, bacteria that synthesize a sialyl alpha 2,8-2,9-linked polymer. A K1- mutant with an insertion mutation in neuS was complemented in trans by K92 neuS, providing direct evidence that neuS encoded the PSA polymerase. A 2.9-kb E. coli K1 kps subclone was sequenced to better characterize polyST. In addition to neuS, the results identified a new open reading frame, designated neuE, the linker sequence between regions 1 and 2, and the last gene of region 1, kpsS. The kpsS translational reading frame was confirmed by sequencing across the junction of a kpsS'-lacZ+ fusion. PolyST was identified by maxicell analysis of nested deletions and coupled in vitro transcription-translation assays. PolyST's derived primary structure predicted a 47,500-Da basic polypeptide without extensive similarity to other known proteins. PolyST activity was increased 31-fold and was membrane localized when neuS was cloned into an inducible expression vector, suggesting, together with the polyST primary structure, that polyST is a peripheral inner membrane glycosyltransferase. However, polyST could not initiate de novo PSA synthesis, indicating a functional requirement for other kps gene products. The existence of a sialyltransferase distinct from polyST was suggested by identification of a potential polyprenyl-binding motif in a C-terminal membrane-spanning domain of the predicted neuE gene product. Direct evidence for a quantitatively minor sialyltransferase activity, which could function to initiate PSA synthesis, was obtained by phenotypic analysis of mutants with multiple defects in sialic acid synthesis, degradation, and polymerization. The results provide an initial molecular description of K1 and K92 sialyltransferase complexes and suggest a possible common function for accessory kps gene products.
Authors:
S M Steenbergen; T J Wrona; E R Vimr
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Publication Detail:
Type:  Journal Article; Research Support, U.S. Gov't, P.H.S.    
Journal Detail:
Title:  Journal of bacteriology     Volume:  174     ISSN:  0021-9193     ISO Abbreviation:  J. Bacteriol.     Publication Date:  1992 Feb 
Date Detail:
Created Date:  1992-03-10     Completed Date:  1992-03-10     Revised Date:  2009-11-18    
Medline Journal Info:
Nlm Unique ID:  2985120R     Medline TA:  J Bacteriol     Country:  UNITED STATES    
Other Details:
Languages:  eng     Pagination:  1099-108     Citation Subset:  IM    
Affiliation:
Department of Pathobiology, College of Veterinary Medicine, University of Illinois, Urbana 61801.
Data Bank Information
Bank Name/Acc. No.:
GENBANK/M76370;  M92996;  M92997;  M92998;  M92999;  M93000;  M93001;  M93002;  M93003;  X56583
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MeSH Terms
Descriptor/Qualifier:
Amino Acid Sequence
Bacterial Capsules / genetics,  metabolism*
Base Sequence
Blotting, Southern
Cloning, Molecular
Escherichia coli / enzymology*,  genetics
Kinetics
Molecular Sequence Data
Plasmids / genetics
Polysaccharides, Bacterial / biosynthesis*
Promoter Regions, Genetic / genetics
Sequence Homology, Nucleic Acid
Sialic Acids / biosynthesis*
Sialyltransferases / chemistry,  genetics,  metabolism*
Grant Support
ID/Acronym/Agency:
AI23039/AI/NIAID NIH HHS
Chemical
Reg. No./Substance:
0/Bacterial Capsules; 0/Polysaccharides, Bacterial; 0/Sialic Acids; 0/polysialic acid; EC 2.4.99.-/Sialyltransferases; EC 2.4.99.1/beta-D-galactoside alpha 2-6-sialyltransferase
Comments/Corrections

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