A Chimeric Nucleotide-Binding Protein, Encoded by a hisP-malK Hybrid Gene, Is Functional in Maltose Transport in Salmonella Typhimurium

Mol Microbiol. 1991 Jun;5(6):1375-83. doi: 10.1111/j.1365-2958.1991.tb00784.x.

Abstract

We have isolated a hybrid gene, composed of the first 455 nucleotides of hisP and nucleotides 275-1107 of malK, the genes coding for the nucleotide-binding components of the high-affinity transport systems for histidine and maltose in Salmonella typhimurium, respectively. The fusion had occurred by recombination within 11 homologous base pairs located between the two DNA fragments. In the chimeric protein peptidic motifs A and B, proposed to be part of the nucleotide-binding fold, originate from HisP and MalK, respectively. Plasmid pES42-39, harbouring the hybrid gene, was shown to complement only a malK mutation but failed to complement a hisP deletion mutation. The chimeric protein was identified by immunoblotting as a protein with an apparent molecular mass of 49kDa. Removal of the C-terminal 77 amino acid residues from the chimeric protein resulted in the loss of function in transport. In contrast, 51 amino acid residues could be removed from the C-terminus of wild-type MalK without any effect. Upon overproduction the chimeric protein, as wild-type MalK, inhibited expression of the malB regulon. However, both truncated proteins, when overproduced, did not exhibit this activity. Based on these results, a tentative model of the functional domains of MalK is presented.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • ATP-Binding Cassette Transporters*
  • Amino Acid Sequence
  • Amino Acid Transport Systems, Basic*
  • Bacterial Proteins / genetics*
  • Bacterial Proteins / metabolism
  • Base Sequence
  • Binding Sites
  • Biological Transport, Active / genetics
  • Carrier Proteins / genetics*
  • Carrier Proteins / metabolism
  • Genetic Complementation Test
  • Histidine / metabolism
  • Immunoblotting
  • Kinetics
  • Maltose / metabolism*
  • Membrane Proteins / genetics
  • Membrane Proteins / metabolism
  • Membrane Transport Proteins / genetics*
  • Membrane Transport Proteins / metabolism
  • Molecular Sequence Data
  • Recombinant Fusion Proteins / genetics*
  • Recombinant Fusion Proteins / metabolism
  • Salmonella typhimurium / genetics
  • Salmonella typhimurium / metabolism*

Substances

  • ATP-Binding Cassette Transporters
  • Amino Acid Transport Systems, Basic
  • Bacterial Proteins
  • Carrier Proteins
  • MalK protein, Bacteria
  • MalK protein, Salmonella typhimurium
  • Membrane Proteins
  • Membrane Transport Proteins
  • Recombinant Fusion Proteins
  • Histidine
  • Maltose
  • histidine permease, Bacteria