Cloning and characterization of a gene, mpsA, encoding a protein associated with intracellular magnetic particles from Magnetospirillum sp strain AMB-1

Proteins located within the Lipid bilayer, surrounding the intracellular ba cterial magnetic particles (BMP) from Magnetospirillum sp. AMB-1, were sepa rated using SDS-PAGE. Several major proteins of approximate molecular weigh t 66.2, 35.6, and 24.8 kDa were identified. The N-terminal amino acid seque nce of one of these proteins, designated MpsA, was determined and used to d esign a pair of PCR primers which amplified a 105 bp DNA fragment from AMB- 1 genomic DNA, Gene-walking, using anchored PCR, was used to determine the complete nucleotide sequence (954 bp) of the mpsA gene. The npsA encodes a 317 amino acid protein which does not have an N-terminal cytoplasmic transp ort signal sequence. Intracellular localization studies were carried out us ing an mpsA-Luc gene fusion expressed in AMB-1 following gene transfer by c onjugation. The gene fusion was constructed by cloning a 1.6 kb mpsA fragme nt upstream of luc in the conjugal plasmid pKLC, The MpsA-Luc fusion protei n was preferentially located on the magnetic particle membrane. Although th e function of MpsA remains unknown, homology searches suggest similarity wi th the cu subunit of acetyl-CoA carboxylase and the CoA-binding motif. (C) 2000 Academic Press.