Genetic Organization of the Cellulose Synthase Operon in Acetobacter xylinum

An operon encoding four proteins required for bacterial cellulose biosynthesis (bcs) in Acetobacter xylinum was isolated via genetic complementation with strains lacking cellulose synthase activity. Nucleotide sequence analysis indicated that the cellulose synthase operon is 9217 base pairs long and...

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Published in:	Proceedings of the National Academy of Sciences - PNAS Vol. 87; no. 20; pp. 8130 - 8134
Main Authors:	Wong, Hing C., Fear, Anna Lisa, Calhoon, Roger D., Eichinger, Gregory H., Mayer, Raphael, Amikam, Dorit, Benziman, Moshe, Gelfand, David H., Measde, James H., Emerick, Anne W., Bruner, Robert, Ben-Bassat, Arie, Tal, Rony
Format:	Journal Article
Language:	English
Published:	Washington, DC National Academy of Sciences of the United States of America 01-10-1990 National Acad Sciences
Subjects:	Acetobacter xylinum Amino acids Arabidopsis Proteins Bacteriology Base Sequence Biological and medical sciences Cloning, Molecular Cosmids DNA Escherichia coli - genetics Fundamental and applied biological sciences. Psychology Genes Genes, Bacterial Genetics Gluconacetobacter xylinus - enzymology Gluconacetobacter xylinus - genetics Glucosyltransferases - genetics Glucosyltransferases - isolation & purification Macromolecular Substances Microbiology Molecular Sequence Data Mutagenesis, Site-Directed Nucleotide sequences Oligonucleotide Probes Operator regions Operon Operons Plasmids Promoter regions Promoter Regions, Genetic Proteins Restriction Mapping Enzymatic activity Gene Mutagenesis Nucleotide sequence Operon Bacteria Gene library Molecular cloning Acetobacter Cellulose synthase (UDP-forming)
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Summary:	An operon encoding four proteins required for bacterial cellulose biosynthesis (bcs) in Acetobacter xylinum was isolated via genetic complementation with strains lacking cellulose synthase activity. Nucleotide sequence analysis indicated that the cellulose synthase operon is 9217 base pairs long and consists of four genes. The four genes--bcsA, bcsB, bcsC, and bcsD--appear to be translationally coupled and transcribed as a polycistronic mRNA with an initiation site 97 bases upstream of the coding region of the first gene (bcsA) in the operon. Results from genetic complementation tests and gene disruption analysis demonstrate that all four genes in the operon are required for maximal bacterial cellulose synthesis in A. xylinum. The calculated molecular masses of the proteins encoded by bcsA, bcsB, bcsC, and bcsD are 84.4, 85.3, 141.0, and 17.3 kDa, respectively. The second gene in the operon (bcsB) encodes the catalytic subunit of cellulose synthase. The functions of the bcsA, bcsC, and bcsD gene products are unknown. Bacterial strains mutated in the bcsA locus were found to be deficient in cellulose synthesis due to the lack of cellulose synthase and diguanylate cyclase activities. Mutants in the bcsC and bcsD genes were impaired in cellulose production in vivo, even though they had the capacity to make all the necessary metabolic precursors and cyclic diguanylic acid, the activator of cellulose synthase, and exhibit cellulose synthase activity in vitro. When the entire operon was present on a multicopy plasmid in the bacterial cell, both cellulose synthase activity and cellulose biosynthesis increased. When the promoter of the cellulose synthase operon was replaced on the chromosome by E. coli tac or lac promoters, cellulose production was reduced in parallel with decreased cellulose synthase activity. These observations suggest that the expression of the bcs operon is rate-limiting for cellulose synthesis in A. xylinum.
Bibliography:	ObjectType-Article-1 SourceType-Scholarly Journals-1 ObjectType-Feature-2 content type line 23
ISSN:	0027-8424 1091-6490
DOI:	10.1073/pnas.87.20.8130