Binding-protein-dependent lactose transport in Agrobacterium radiobacter.
Agrobacterium radiobacter NCIB 11883 was grown in lactose-limited continuous culture at a dilution rate of 0.045/h. Washed cells transported [14C]lactose and [methyl-14C]beta-D-thiogalactoside, a nonmetabolisable analog of lactose, at similar rates and with similar affinities (Km for transport, less than 1 microM). Transport was inhibited to various extents by the uncoupling agent carbonyl cyanide p-trifluoromethoxyphenylhydrazone, by unlabeled beta-galactosides and D-galactose, and by osmotic shock. The accumulation ratio for methyl-beta-D-thiogalactoside was greater than or equal to 4,100. An abundant protein (molecular weight, 41,000) was purified from osmotic-shock fluid and shown by equilibrium dialysis to bind lactose and methyl-beta-D-thiogalactoside, the former with very high affinity (binding constant, 0.14 microM). The N-terminal amino acid sequence of this lactose-binding protein exhibited some homology with several other sugar-binding proteins from bacteria. Antiserum raised against the lactose-binding protein did not cross-react with two glucose-binding proteins from A. radiobacter or with extracts of other bacteria grown under lactose limitation. Lactose transport and beta-galactosidase were induced in batch cultures by lactose, melibiose [O-alpha-D-galactoside-(1----6)alpha-D-glucose], and isopropyl-beta-D-thiogalactoside and were subject to catabolite repression by glucose, galactose, and succinate which was not alleviated by cyclic AMP. We conclude that lactose is transported into A. radiobacter via a binding protein-dependent active transport system (in contrast to the H+ symport and phosphotransferase systems found in other bacteria) and that the expression of this transport system is closely linked to that of beta-galactosidase.
Greenwood
J A
JA
Department of Biochemistry, University of Leicester, United Kingdom.
Cornish
A
A
Jones
C W
CW
D000595:Amino Acid Sequence; D001693:Biological Transport, Active; D029968:Escherichia coli Proteins; D007700:Kinetics; D007785:Lactose / Q000378:metabolism*; D026901:Membrane Transport Proteins / Q000235:genetics / Q000378:metabolism*; D008756:Methylgalactosides / Q000378:metabolism; D008969:Molecular Sequence Data; D008970:Molecular Weight; D009004:Monosaccharide Transport Proteins; D012231:Rhizobium / Q000201:enzymology* / Q000235:genetics; D012689:Sequence Homology, Nucleic Acid; D027981:Symporters; D013862:Thiogalactosides / Q000378:metabolism
D029968:Escherichia coli Proteins; C050221:LacY protein, E coli; D026901:Membrane Transport Proteins; D008756:Methylgalactosides; D009004:Monosaccharide Transport Proteins; D027981:Symporters; D013862:Thiogalactosides; C009468:thiomethylgalactoside; C021499:lactose permease; D007785:Lactose