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The outer membrane of Vibrio cholerae contains a maltose-inducible major protein, OmpS (43 kDa), that is common to different isolates. Nucleotide sequence analysis of the corresponding structural gene, ompS, revealed an open reading frame encoding a 412-amino-acid polypeptide. The amino acid sequence of OmpS is similar to that of LamB, the Escherichia coli maltoporin, and to ScrY of Kiebsiella pneumoniae, although the antigenic determinants of these proteins are different. The cloned ompS gene complemented an ompS mutation of V. cholerae and the corresponding polypeptide could function as a maltoporin in a LamB- mutant of E. coli The promoter region of ompS is highly homologous to the malK-lamB promoter of E. coli and the ompS gene is controlled by MalT in E. coli This indicates that the same kind of regulatory mechanism is used to activate the ompS expression in V. cholerae and malK-lamB expression in E. coli An ompS-lacZ transcriptional fusion was used to demonstrate a dual control in ompS expression; the ompS gene is responsive to the inducers maltose and trehalose but in their absence it is also expressed in response to growth-phase. These different modes of induction might be of importance during different stages of V. cholerae infection. [ABSTRACT FROM AUTHOR]

The maIG gene encodes a hydrophobic cytoplasmic membrane protein which is required for the energy-dependent transport of maltose and maltodextrins in Escherichia coli. The MaIG protein, together with MalF and MalK proteins, forms a multimeric complex in the membrane consisting of two MalK subunits for each MalF and MaIG subunit. Fifteen mutations have been isolated in maIG by random linker insertion mutagenesis. Two regions essential for maltose transport have been identified. In particular, a hydrophjiic region containing the peptidic motif EAA-G---I-LP, highly conserved among inner membrane proteins from binding protein-dependent transport systems, is essential for maltose transport. The results also show that several regions of MaIG are not essential for function. A region (residues 30-50) encompassing the first predicted transmembrane segment and the first periplasmic loop in MaIG may be modified extensively with little effect on maltose transport and no effect on the stability and the localization of the protein. A region located at the middle of the protein (residues 153-157) is not essential for the function of the protein. A region, essential for maltodextrin utilization but not for maltose transport, has been identified near the C-terminus of the protein. [ABSTRACT FROM AUTHOR]

The genes coding for the binding-protein-dependent lactose transport system and β-galactosidase in Agrobacterium radiobacter strain AR50 were cloned and partially sequenced. A novel lac operon was Identified which contains genes coding for a lactose-binding protein (lacE), two integral membrane proteins (lacF and lacG), an ATP-binding protein (lacK) and β-galactosidase (lacZ). The operon is transcribed in the order lacEFGZK. The operon is controlled by an upstream regulatory region containing putative -35 and -10 promoter sites, an operator site, a CRP-binding site probably mediating catabolite repression by glucose and galactose, and a regulatory gene (lacl) encoding a repressor protein which mediates induction by lactose and other galactosides in wild-type A. radiobacter (but not in strain AR50, thus allowing constitutive expression of the lac operon). The derived amino acid sequences of the gene products indicate marked similarities with other binding-protein-dependent transport systems in bacteria. [ABSTRACT FROM AUTHOR]

During the molecular analysis of a plasmid-coded sucrose metabolic pathway of enteric bacteria, a gene, scrY, was found whose product, ScrY, had all the properties of a bacterial porin (Schmid et al., 1988). Loss of this protein (Mr, 58kDa), localized in the outer membrane, led, as shown here, to an increase in the apparent Km, for sucrose transport in whole cells from 10µM in wild-type cells to 300µM in mutant cells. This contrasts with the Km for sucrose phosphorylation as measured in membrane vesicles from mutant and wild-type cells, which remained unchanged at about 10 µM, and reflects the activity of the sucrose-specific Enzymell of the phosphoenolpyruvate-dependent carbohydrate:phosphotransf erase system (PTS) responsible for uptake through the inner membrane. Furthermore, the presence of ScrY restored growth on maltodextrins in cells devoid of LamB, thus complementing the lack of this maltoporin. The amino acid sequence deduced from the DNA sequence was determined for the plasmid-coded and the ScrY porin coded in the chromosome of Klebsiella pneumoniae. Both show high identity (86%) to each other, and to the channel domain of LamB, further corroborating the conclusion that they constitute porins. [ABSTRACT FROM AUTHOR]

The oprP gene encoding the Pseudomonas aeruginosa phosphate-specific outer membrane porin protein OprP was sequenced. Comparison of the derived amino acid sequence with the known sequences of other bacterial porins demonstrated that OprP could be no better aligned to these porin sequences than it could to the periplasmic phosphate-binding protein PhoS of Escherichia coli. Southern hybridization and restriction mapping of the oprP gene in 37 clinical isolates and the 17 serotype strains of P. aeruginosa revealed that restriction sites in the vicinity of the oprP gene were highly conserved. Several species from the Pseudomonas fluorescens rRNA homology group contained DNA that hybridized to an oprP gene probe. [ABSTRACT FROM AUTHOR]

The nucleotide sequence of the ugp genes of Escherichia coli K-12, which encode a phosphate-limitation inducible uptake system for sn-glycerol-3-phosphate and glycerophosphoryl diesters, was determined. The genetic organization of the operon differed from previously published results. A single promoter, containing a putative pho box, was detected by S1-nuclease mapping. The promoter is followed by four open reading frames, designated ugpB, A, E and C, which encode a periplasmic binding protein, two hydrophobic membrane proteins and a protein that is likely to couple energy to the transport system, respectively. The sequences of the proteins contain the characteristics of several other binding protein-dependent transport systems, but they seem to be particularly closely related to the maltose system. [ABSTRACT FROM AUTHOR]

Presents the genetics of active transport in bacteria. Inclusion of the fundamental processes of all cells; Location of functionally important sites for substrate recognition and energy coupling; Conformational changes that occur during the transfer of substrate from one side of the membrane to the other.

The kinetic parameters for the maltose transport system in Escherichia coli K12 were determined with maltose and maltotriose as substrates. The system exhibits an apparent Km of 1μM for maltose and 2 μM for maltotriose. The V of entry was determined as 2.0 and 1.1 mol substrate/min per 108 cells. Mutations in lamB, the structural gene for the receptor protein of phage λ, increased the Km for maltose transport by a factor of 100–500 without influencing the maximal rate of transport. Maltotriose is no longer transported in these lamB mutants. The maltose-binding protein, an essential component of the maltose transport system, was found .to exhibit substrate-dependent fluorescence quenching. This phenomenon was used to determine dissociation constants and to estimate the rate of ligand dissociation. A Kd of 1 μM for maltose and of 0.16 μM for maltotriose was found. From the comparison of the kinetic parameters of transport of maltose and maltotriose in wild-type and λ-resistant mutants with the binding constants for both sugars to purified maltose-binding protein, we conclude that the λ receptor facilitates the diffusion of maltose and maltodextrins through the outer membrane. [ABSTRACT FROM AUTHOR]

In Escherichia coli K12 the product of gene lamB is an outer membrane protein involved in the transport of maltose and maltodextrins and serving as a receptor for several bacteriophages including λ. About 30 to 40% of this protein can be recovered associated to peptidoglycan when the cells are dissolved in sodium dodecyl sulfate in the presence of 2 mM Mg2+ ions. The bound protein can then be quantitatively eluted from peptidoglycan by incubating the complex in Triton X-100 and EDTA, or sodium dodecyl sulfate and NaCl. The protein eluted in such ways is still totally active in its phage-neutralizing activity. Two other outer membrane proteins known to behave similarly to the lamB protein are proteins Ia and lb. However the binding of these proteins to peptidoglycan appears tighter, in several respects, than that of the lamB protein. The lamB protein may span the outer membrane since it appears to interact with the peptidoglycan on the inner side of this membrane while it is known to be accessible to both phages and antibodies at the cell surface. [ABSTRACT FROM AUTHOR]