Reduction of methylglyoxal in Escherichia coli K12 by an aldehyde reductase and alcohol dehydrogenase

Two enzymes, one NADPH-dependent and another NADH-dependent which catalyze the reduction of methylglyoxal to acetol have been isolated and substantially purified from crude extracts of Escherichia coli K₁₂ cells. Substrate specificity and formation of acetol as the reaction product by both the enzymes, reversibility of NADH-dependent enzyme with alcohols as substrates and inhibitor study with NADPH-dependent enzyme indicate that NADPH-dependent and NADH-dependent enzymes are identical with an aldehyde reductase (EC 1.1.1.2) and alcohol dehydrogenase (EC 1.1.1.1) respectively. The K_m for methylglyoxal have been determined to be 0.77 mM for NADPH-dependent and 3.8 mM for NADH-dependent enzyme. Stoichiometrically equimolar amount of acetol is formed from methylglyoxal by both NADPH- and NADH-dependent enzymes. In phosphate buffer, both the enzymes are active in the pH range of 5.8–6.6 with no sharp pH optimum. Molecular weight of both the enzymes were found to be 100,000 ± 3,000 by gel filtration on a Sephacryl S-200 column. Both NADPH- and NADH-dependent enzymes are sensitive to sulfhydryl group reagents.     

Interactions ofCandida albicans with bacteria and salivary molecules in oral biofilms

The yeastCandida albicans coaggregates with a variety of streptococcal species, an interaction that may promote oral colonization by yeast cells.C. albicans andCandida tropicalis are the yeasts most frequently isolated from the human oral cavity and our data demonstrate that both these species bind toStreptococcus gordonii NCTC 7869 while two otherCandida species (Candida krusei andCandida kefyr) do not. Adherence ofC. albicans was greatest when the yeast had been grown at 30° C to mid-exponential growth phase. For 21 strains ofC. albicans there was a positive correlation between the ability to adhere toS. gordonii and adherence to experimental salivary pellicle. Whole saliva either stimulated or slightly inhibited adherence ofC. albicans toS. gordonii depending on the streptococcal growth conditions. The results suggest that the major salivary adhesins and coaggregation adhesins ofC. albicans are co-expressed.     

Changes in UsnRNA biosynthesis during rat liver regeneration

Partial hepatectomy (P.H.) induces a partially synchronized growth response of liver under normal regulation of growth. In this phase changes in cellular morphology, radial distribution pattern of cells and other biological as well as major biochemical changes are well documented [24]. Here, we have shown that the cellular content of UsnRNAs altered during this proliferative phase as well. The level of spliceosomal UsnRNAs (U1, U2, U4–U6) gradually decreased by 30–50% upto 48 hrs of P.H. followed by gradual increase to reach the normal level within one month of P.H. The U3 snRNA level on the other hand, was nearly equal to that in normal liver at 48 hrs of P.H. but in 24 and 72 hrs of P.H. its level was high (4 fold) in contrast to that in other UsnRNAs. Thus, it is clear from our data that the level of all the six UsnRNAs decreased during 48 hrs of P.H. compared to that after first 24 hrs. This has been correlated in the kinetics of UsnRNAs' synthesis (in terms of labelling) in isolated hepatocytes, where the rate of labelling of all the six UsnRNAs increased 20–30% in 24 hrs regenerating hepatocytes (R.H.) followed by sharp decrease by 30–50% within next 24 hrs, compared to that in the normal hepatocytes. But from 72 hrs onwards in R.H. the rate of labelling of all the six UsnRNAs again increased by 30–50% (compared to that in normal hepatocytes) followed by decrease of their labelling-rate to reach the normal level in R.H. within one month of P.H. Thus, it may be concluded that the changes in UsnRNAs' level during the proliferative phase of liver regeneration may be either due to the alteration in the rate of synthesis (in terms of labelling) or along with it differential turn over rate; this phenomenon may have some consequences with the regenerative process of liver.This paper was published in Molecular and Cellular Biochemistry131:67–73, 1994. Kluwer Academic Publishers regret the publication of the only partly corrected version.     

Effects of posture, hypotension and locally applied vasoconstriction on the middle ear microcirculation in anaesthetized humans

Studies by laser-Doppler flowmetry of middle ear microcirculation changes induced by physical and chemical stimuli in the animal have only recently been made. This prospective study, performed in humans, was designed to compare the effects of a postural manoeuvre (headup tilt 30°), hypotension and locally applied vasoconstriction on middle ear blood flow during anaesthesia. Circulatory changes provoked by a headup tilt of 30°, and successive intravenous boluses of potent vasodilators, were compared with circulatory changes provoked by locally applied adrenaline, in ten healthy patients in good physical states undergoing middle ear surgical repair. Heart rate and direct arterial pressure were continuously recorded via a radial artery cannula. Middle ear blood flow was continuously recorded via a laser-Doppler probe placed on the promontorium cavi tympani. Metabolic parameters (partial pressure of O₂ and CO₂ in arterial blood, pH, arterial lactate concentrations) and arterial concentrations of propofol were measured just before and just after the experiment. Headup tilt did not modify heart rate, mean arterial pressure or middle ear blood flow. Vasodilators (nicardipine, nitroprusside, nitroglycerin) provoked a fall in arterial pressure (P<0.0001,P<0.0001,P<0.019, respectively), but did not induce any significant variations in heart rate; variations occurred in middle ear blood flow (P>0.05, not significant) which were different according to patients and agents. Locally applied adrenaline provoked a fall in the middle ear blood flow (P<0.0012), with no effect on heart rate and arterial pressure. There were no significant changes in metabolic values, or propofol serum concentrations. The behaviour of the middle ear blood flow submitted to hypotension, posture, or to vasoconstriction could be related to counteracting regulatory responses and/or to direct vascular effects.     

β-Amylase fromBacillus megaterium

Bacillus megaterium strain B₆ producing extracellular β-amylase was isolated and grown in a medium supplemented with waste potato starch. It showed highest enzyme synthesis in the early stationary phase. The partially purified β-amylase had a temperature optimum at 60°C and a pH optimum at 6.9 and was not affected by Schardinger dextrins. These properties would allow its application in sugar industry.     

Phosphorylation of lipopolysaccharides in the Antarctic psychrotroph Pseudomonas syringae: a possible role in temperature adaptation.

Phosphorylation of lipopolysaccharide (LPS) from a psychrotrophic bacterium, Pseudomonas syringae, from Antarctica was studied by using sucrose gradient-separated membrane fractions. The bacterium was found to possess an LPS kinase which could phosphorylate more LPS postsynthetically at higher temperatures. The phosphorylation was low at a lower temperature and was also found to occur in vivo. After phosphorylation of LPS in vitro, it was found that the major part of the radioactivity (> 85%) was associated with the core oligosaccharide region of the LPS. The phosphate groups of this region are probably involved in the binding of metal ions, which could be removed by EDTA. The cells grown at the lower temperature probably contained fewer divalent cations because of the smaller amount of phosphate and thereby were more sensitive to EDTA. The cells were also more sensitive to cationic antibiotics at the lower temperature. A possible role of this differential phosphorylation of LPS in modulating the function of the outer membrane as a permeability barrier in the psychrotroph is discussed.     

Fibroblasts isolated from human pterygia exhibit transformed cell characteristics

Summary Pterygium is a degenerative corneal limbal process and UV irradiation has been suggested as being a major environmental predisposing factor. The invasive nature of the fibroblasts associated with pterygia raises the question as to whether these cells are transformed. To test this hypothesis, we established fibroblast strains from autologous and heterologous pterygial and conjunctival specimens, respectively, from subjects between 40 to 50 yr of age, and compared their growth characteristics in culture. All pterygial fibroblast strains exhibited a reduced dependence on serum and exogenous growth factors for growth and reached a saturation population density that was threefold higher than conjunctival fibroblasts cultured under the same conditions. In addition, all pterygial fibroblast strains were able to form colonies in soft agar in 5% fetal bovine serum at a 6.0 to 7.5% efficiency. Under the same experimental conditions, none of the conjunctival fibroblast strains were able to grow. The results presented support the conclusion that pterygial fibroblasts have acquired many of the properties of the transformed phenotype.