RESUMO
Listeria monocytogenes is a food-borne pathogen with a high mortality rate that has also emerged as a paradigm for intracellular parasitism. We present and compare the genome sequences of L. monocytogenes (2,944,528 base pairs) and a nonpathogenic species, L. innocua (3,011,209 base pairs). We found a large number of predicted genes encoding surface and secreted proteins, transporters, and transcriptional regulators, consistent with the ability of both species to adapt to diverse environments. The presence of 270 L. monocytogenes and 149 L. innocua strain-specific genes (clustered in 100 and 63 islets, respectively) suggests that virulence in Listeria results from multiple gene acquisition and deletion events.
Assuntos
Proteínas de Bactérias/genética , Genoma Bacteriano , Listeria monocytogenes/genética , Listeria/genética , Adaptação Fisiológica , Motivos de Aminoácidos , Bacillus subtilis/genética , Proteínas de Bactérias/química , Proteínas de Bactérias/fisiologia , Composição de Bases , Proteínas de Transporte/química , Proteínas de Transporte/genética , Cromossomos Bacterianos/genética , DNA Bacteriano/química , DNA Bacteriano/genética , Transferência Genética Horizontal , Genes Bacterianos , Genômica , Listeria/química , Listeria/fisiologia , Listeria monocytogenes/química , Listeria monocytogenes/patogenicidade , Listeria monocytogenes/fisiologia , Proteínas de Membrana/química , Proteínas de Membrana/genética , Análise de Sequência de DNA , Staphylococcus aureus/genética , Fatores de Transcrição/química , Fatores de Transcrição/genética , Virulência/genéticaRESUMO
Sequence comparisons of members of the myosin superfamily show a high degree of charge conservation in a surface exposed helix (Dictyostelium discoideum myosin II heavy chain residues S510 to K546). Most myosins display a triplet of acidic residues at the equivalent positions to D. discoideummyosin II residues D530, E531, and Q532. The high degree of charge conservation suggests strong evolutionary constrain and that this region is important for myosin function. Mutations at position E531 were shown to strongly affect actin binding [Giese, K. C., and Spudich, J. A. (1997) Biochemistry 36, 8465-8473]. Here, we used steady-state and transient kinetics to characterize the enzymatic competence of mutant constructs E531Q and Q532E, and their properties were compared with those of a loop 2 mutant with a 20 amino acid insertion containing 12 positive charges (20/+12) [Furch et al. (1998) Biochemistry 37, 6317-6326], double mutant Q532E(20/+12), and the native motor domain constructs. Our results confirm that charge changes at residues 531 and 532 primarily affect actin binding with little change being communicated to the nucleotide pocket. Mutation D531Q reduces actin affinity (K(A)) 10-fold, while Q532E leads to a 5-fold increase. The observed changes in K(A)() stem almost exclusively from variations in the dissociation rate constant (k(-A)), with the introduction of a single negative charge at position 532 having the same effect on k(-A) as the introduction of 12 positive charges in the loop 2 region.