Summary Listeria monocytogenes is a Gram

Salmonella enterica sv. typhimurium (S. enterica sv. Typhimurium) has two metal-transporting P1-type ATPases whose actions largely overlap with respect to growth in elevated copper. Mutants lacking both ATPases over-accumulate copper relative to wild-type or either single mutant. Such duplication of ATPases is unusual in bacterial copper tolerance. Both ATPases are under the control of MerR family metal-responsive transcriptional activators. Analyses of periplasmic copper complexes identified copper-CueP as one of the predominant metal pools. Expression of cueP was recently shown to be controlled by the same metal-responsive activator as one of the P1-type ATPase genes (copA), and copper-CueP is a further atypical feature of copper homeostasis in S. enterica sv. Typhimurium. Elevated copper is detected by a reporter construct driven by the promoter of copA in wild-type S. enterica sv. Typhimurium during infection of macrophages. Double mutants missing both ATPases also show reduced survival inside cultured macrophages. It is hypothesized that elevated copper within macrophages may have selected for specialized copper-resistance systems in pathogenic microorganism such as S. enterica sv. Typhimurium.

Monoclonal antibodies, induced with a phosphonate diester hapten, catalyzed the coupling of p-nitrophenyl esters of N-acetyl valine, leucine, and phenylalanine with tryptophan amide to form the corresponding dipeptides. All possible stereoisomeric combinations of the ester and amide substrates were coupled at comparable rates. The antibodies did not catalyze the hydrolysis of the dipeptide product nor hydrolysis or racemization of the activated esters. The yields of the dipeptides ranged from 44 to 94 percent. The antibodies were capable of multiple turnovers at rates that exceeded the rate of spontaneous ester hydrolysis. This achievement suggests routes toward creating a small number of antibody catalysts for polypeptide syntheses.

We describe here the identification and characterization of two Listeria monocytogenes (Tn917-LTV3) relA and hpt transposon insertion mutants that were impaired in growth after attachment to a model surface. Both mutants were unable to accumulate (p)ppGpp in response to amino acid starvation, whereas the wild-type strain accumulated (p)ppGpp within 30 min of stress induction. The induction of transcription of the relA gene after adhesion was demonstrated, suggesting that the ability to mount a stringent response and undergo physiological adaptation to nutrient deprivation is essential for the subsequent growth of the adhered bacteria. The absence of (p)ppGpp in the hpt mutant, which is blocked in the purine salvage pathway, is curious and suggests that a functional purine salvage pathway is required for the biosynthesis of (p)ppGpp. Both mutants were avirulent in a murine model of listeriosis, indicating an essential role for the stringent response in the survival and growth of L. monocytogenes in the host. Taken as a whole, this study provides new information on the role of the stringent response and the physiological adaptation of L. monocytogenes for biofilm growth and pathogenesis.The ubiquitous gram-positive pathogen L. monocytogenes is responsible for the clinical syndromes of listeriosis in humans and animals (27, 28). The principle mode of transmission of the organism to humans is believed to be the consumption of contaminated food (44). Normally, the consumption of foodborne L. monocytogenes does not result in overt gastrointestinal disease. Rather, in serious cases, listeriosis presents as bacteremia, meningitis, or miscarriage (30).The ability of L. monocytogenes to adhere to and colonize surfaces during food preparation and storage is important in the contamination of food products prior to consumption. While much is known about the virulence of L. monocytogenes (reviewed in references 7 and 24), detailed knowledge about the mechanisms of attachment and proliferation of L. monocytogenes on surfaces is scant. Previously, the adhesion of L. monocytogenes to plant surfaces has been shown to be a consequence of hydrophobic bonds between the plant surface and outer surface components of the bacterium rather than a specific ligand-receptor binding process (3).It is known that biofilm formation is a complex process, and a number of cell surface structures have been implicated in the initial stages of biofilm formation. In Pseudomonas aeruginosa these include flagella (motility) (35) and the expression of type IV pili for microcolony formation (36). In streptococci a number of cell surface adhesins, together with autolysins, have been identified as being important in biofilm development (12,17,19,21,26). The subsequent development and differentiation of an ordered three-dimensional biofilm requires the expression of extracellular polysaccharides (9) and may involve cell-to-cell signaling molecules (11,26). During the initial establishment of a sessile community, it is likely that bacteria have to adapt their...

Heparin remains a major drug in prevention of thromboembolic disease. Concerns related to its animal source have prompted search for heparin analogues. The anticoagulant activity of heparin depends on a specific pentasaccharide sequence that binds antithrombin. We report the generation of a product with antithrombin-binding, anticoagulant, and antithrombotic properties similar to those of heparin, through combined chemical and enzymatic modification of a bacterial (E. coli K5) polysaccharide. The process is readily applicable to large-scale production.