A 71-KDA PROTEIN FROM HALOBACTERIUM-SALINARIUM BELONGS TO A UBIQUITOUS P-LOOP ATPASE SUPERFAMILY WITH HEAD-ROD-TAIL STRUCTURE

The nucleotide sequence of a genomic fragment from Halobacterium salin arium containing an open reading frame encoding a protein with a calcu lated molecular mass of 71 kDa was determined. Database searches revea led that this protein, Hp71, has similarities to eukaryotic cytoskelet al proteins. Heterologous production of Hp71 in Escherichia coil allow ed the isolation of anti-Hp71 antibodies. The antibodies were used (1) to verify the production of Hp71 in H. salinarium and (2) to determin e its cytoplasmic localization by immune electron microscopy. Homologo us overproduction of Hp71 in H. salinarium and heterologous production in Haloferax volcanii resulted in modifications of cell morphology fr om rods to extended rods, and from pleiomorphic cells to rods, respect ively. Structure prediction methods indicated that Hp71 has a head-rod -tail configuration, including an N-terminal domain with a nucleotide binding motif (P-loop), and an extended discontinuous coiled-coil doma in of 330 amino acids. To identify related proteins, the complete geno mes of Haemophilus influenzae, Mycoplasma genitalium, and Methanococcu s jannaschii were searched for deduced proteins with extended coiled-c oil domains. Only one or two proteins were found for each organism, sh owing that Hp71 is one of only a few prokaryotic intracellular protein s with extended coiled-coil domains. The phenotype upon overproduction and the similarity of Hp71 to the SMC superfamily of P-loop head-rod- tail proteins (named after SMC1, which is involved in the ''stability of minichromosomes'' in yeast) indicate that Hp71 might be involved in cytoskeleton formation and/or chromosome partitioning in H. salinariu m.