Cloning of a gene involved in cellulose biosynthesis in Acetobacter xylinum: complementation of cellulose-negative mutants by the UDPG pyrophosphorylase structural gene.
Three cellulose-negative (Cel-) mutants of Acetobacter xylinum strain ATCC 23768 were complemented by a cloned 2.8 kb DNA fragment from the wild type. Biochemical analysis of the mutants showed that they were deficient in the enzyme uridine 5'-diphosphoglucose (UDPG) pyrophosphorylase. The analysis also showed that the mutants could synthesize beta(1-4)-glucan in vitro from UDPG, but not in vivo from glucose. This result was expected, since UDPG is known to be the precursor for cellulose synthesis in A. xylinum. In order to analyze the function of the cloned gene in more detail, its biological activity in Escherichia coli was studied. These experiments showed that the cloned fragment could be used to complement an E. coli mutant deficient in the structural gene for UDPG pyrophosphorylase. It is therefore clear that the cloned fragment must contain this gene from A. xylinum. This is to our knowledge the first example of the cloning of a gene with a known function in cellulose biosynthesis from any organism, and we suggest the gene be designated celA.
Valla
S
S
Laboratory of Biotechnology, Norwegian Institute of Technology, Trondheim.
Coucheron
D H
DH
Fjaervik
E
E
Kjosbakken
J
J
Weinhouse
H
H
Ross
P
P
Amikam
D
D
Benziman
M
M
D002482:Cellulose / Q000096:biosynthesis / Q000235:genetics; D003001:Cloning, Molecular; D003227:Conjugation, Genetic; D004269:DNA, Bacterial / Q000235:genetics; D004587:Electrophoresis, Agar Gel; D029968:Escherichia coli Proteins; D005796:Genes; D005816:Genetic Complementation Test; D041822:Gluconacetobacter xylinus / Q000201:enzymology / Q000235:genetics*; D008190:Lyases / Q000378:metabolism; D009154:Mutation; D009713:Nucleotidyltransferases / Q000235:genetics*; D010733:Phosphoglucomutase / Q000378:metabolism; D010727:Phosphoric Diester Hydrolases / Q000378:metabolism; D019761:Phosphorus-Oxygen Lyases; D010957:Plasmids; D005957:UTP-Glucose-1-Phosphate Uridylyltransferase / Q000235:genetics*
D004269:DNA, Bacterial; D029968:Escherichia coli Proteins; D002482:Cellulose; D009713:Nucleotidyltransferases; D005957:UTP-Glucose-1-Phosphate Uridylyltransferase; D010727:Phosphoric Diester Hydrolases; D008190:Lyases; D019761:Phosphorus-Oxygen Lyases; C060720:diguanylate cyclase; D010733:Phosphoglucomutase