Pressure-adaptive differences in proteolytic inactivation of M4-lactate dehydrogenase homologues from marine fishes

The inactivation by hydrostatic pressure of muscle-type lactate dehydrogenase (M4-LDH, EC 1.1.1.27; L-lactate: NAD+ oxidoreductase) homologues from five shallow-living and six deep-living marine teleost fishes was compared. The pressures which inactivate these enzymes are much higher than the pressures experienced by any of the species. To determine whether hydrostatic pressure effects on protein aggregation state and conformation might influence proteolysis, the inactivation of LDH by the proteases, trypsin (EC 3.4.21.4) and subtilisin (EC 3.4.4.16) was determined at atmospheric pressure and 1,000 atm pressure. At 10 degrees C and atmospheric pressure, the enzymes of the shallow-living fishes are inactivated four times faster by trypsin and three times faster by subtilisin than are the homologues of the deep-living species. At 1,000 atm pressure, the homologues of shallow-occurring fishes were inactivated 28 to 64% more than predicted from the summed effects of denaturation by 1,000 atm pressure and tryptic inactivation at atmospheric pressure. In contrast, the homologues of the deep-sea species were inactivated by trypsin 0 to 21% more than expected. At 1,000 atm, inactivation by subtilisin increased to a similar degree for enzymes from both deep- and shallow-living species. However, at 1,000 atm, the M4-LDH homologues of the deep-sea species lost less activity (55.3%) than did the homologues of the shallow species (86.4%). In comparisons made at 200 atm, a pressure typical of the habitat of the deep-occurring species, tryptic inactivation of the LDH of the shallow-living Sebastes melanops was increased 14%. No pressure inactivation of the enzyme is evident at 200 atm.(ABSTRACT TRUNCATED AT 250 WORDS)     

Growth of Paramecium tetraurelia in bacterized, monoxenic cultures

Wild type and mutant Paramecium tetraurelia were grown in monoxenic cultures by first growing Enterobacter aerogenes on a defined medium and then adding the Paramecium to the stationary phase bacterial culture. The bacterial growth was proportional to the concentration of the carbon source (citrate), and the Paramecium growth was dependent upon both the bacterial density and the starting density of Paramecium. The behavior, electrophysiological properties, ciliary lipid composition, and growth characteristics were similar to the commonly used bacterized medium (Cerophyl) except that 5-10 times greater Paramecium yields were reliably obtained.     

Regulation of glucosyl- and fructosyltransferase synthesis by continuous cultures of Streptococcus mutans

Streptococcus mutans strains Ingbritt, and its derivative B7 which had been passaged through monkeys, have been used to investigate how the synthesis of extracellular glucosyl- and fructosyltransferases is regulated. The most active enzyme from carbon-limited continuous cultures was a fructosyltransferase; enzymes catalysing the formation of water-insoluble glucans from sucrose were relatively inactive. Dextransucrase (EC 2.4.1.5), which catalyses soluble glucan synthesis, was most active in the supernatant fluid from cultures grown with excess glucose, fructose or sucrose, but full activity was detected only when the enzyme was incubated with both sucrose and dextran. Little dextransucrase activity was detected in carbon-limited cultures. It is concluded that glucosyl- and fructosyltransferases are constitutive enzymes in that they are synthesized at similar rates during growth with an excess of the substrate or of the products of the reactions which they catalyse. Although the Ingbritt strain was originally isolated from a carious lesion, it is now a poor source of glucosyltransferase activity. Glucosyltransferases were extremely active in cultures of a recent clinical isolate, strain 3209, and were apparently induced during growth with excess glucose.