L-serine Dehydratase Activity Research Articles

L-serine catabolism via an oxygen-labile L-serine dehydratase is essential for colonization of the avian gut by Campylobacter jejuni.

Campylobacter jejuni is a microaerophilic, asaccharolytic bacterium. The identity of the carbon and energy sources used by C. jejuni in vivo is unknown, but the genome sequence of strain NCTC11168 indicates the presence of genes for catabolism of a limited range of amino acids, including serine. Specific omission of L-serine from a defined medium containing a mixture of amino acids led to a dramatic decrease in cell yields. As C. jejuni does not have a biosynthetic serine requirement, this supports earlier suggestions that L-serine is a preferentially catabolized amino acid. Serine transport was found to be mediated by at least two systems in strain 11168; a high-capacity, low-affinity L-serine-specific system encoded by Cj1625c (sdaC) and a higher-affinity L-serine/L-threonine system responsible for residual L-serine transport in an sdaC mutant. Catabolism of L-serine to pyruvate and ammonia is carried out by SdaA (encoded by Cj1624c), which was overexpressed, purified, and shown to be an oxygen-labile iron-sulfur enzyme. L-Serine dehydratase activity in an sdaA mutant was reduced 10-fold compared to that in the wild type, but the residual activity (due to the anabolic L-threonine dehydratase) could not support either growth on or utilization of L-serine in defined media. However, although sdaA mutants showed no obvious growth defect in complex media, they completely failed to colonize 3-week-old chickens as assayed both by cloacal swabs taken over a 6-week period and by cecal colony counts postmortem. In contrast, the isogenic parent strain colonized chickens to high levels within 1 week of inoculation. The results show that an active SdaA is essential for colonization of the avian gut by C. jejuni and imply that catabolism of L-serine is crucially important for the growth of this bacterium in vivo.

Effects of l-serine dehydratase activity on l-serine production by Corynebacterium glycinophilum and an examination of the properties of the enzyme

The results with Corynebacterium glycinophilum AJ-3170 and various mutants from AJ-3170 indicated that l-serine production was almost inversely proportional to l-serine degrading activity. The crude extract of the parental strain, AJ-3170, showed l-serine and l-threonine degrading activities. The 2 activities were completely separated from each other by gel-filtration, indicating that each activity comes from a different enzyme. The l-serine degrading enzyme, l-serine dehydratase (SD), was purified 30-fold from AJ-3170. Molecular weight of SD was 130,000. The enzyme was specific for l-serine, activated slightly by FeCl 2 and inhibited by MnCl 2. The double reciprocal plots of SD rate against substrate concentration gave an upwards-curved line. The value of [S] 0.5 was 35 mM.

Characterization of a L-serine dehydratase activity from Streptococcus faecalis

Streptococcus faecalis sp. produces pyruvate and ammonia from L-serine via a specifie L-serine dehydratase. The apparent Michaelis. constant for L-serine is 50 mM and the Vmax is 142 nmol of pyruvate formed per minute and per mg of protein. Maximum enzymatic activity is observed at 40 C and pH 8 in 100 mM phosphate buffer. L-serine is the sole substrate. D-serine, L-threonine and glycine are competitive inhibitors, L-cysteine acts as a non-competitive inhibitor.

Characterization of an L-serine dehydratase activity in permeabilized cells of Brevibacterium linens ATCC 9175

Brevibacterium linens ATCC 9175 praduces large quantities of, ammonia and pyruvate (rom L-serine. This reaction occurs via an L-serine dehydratase (EC.4.2.I.13), whose activity is maximal at the end of exponential growth on rich medium and which abruptly decreases at the beginning of the stationary phase. ft is not possible ta extract the soluble form of the enzyme without a total loss of activity. Subcellular localization studies after creating stable proto- plasts with lysozyme have shawn that a part of the activity is bound ta cell membranes. A stable (orm of activity has been obtained by treating the cells with Triton X-IOO and the main properties of the enzyme have been studied with this insoluble form. The apparent Michaelis constant is 50 mM and the Vmax is 200 nmol of pyruvate formed per minute and per mg of dry extract. The optimum pH is included between 8.5 and 9.4 anâ activity is very stable between 8.0 and 9.4. The enzyme is slightly thermostable. The exclusive substrate is L-serine. D-serine and 2-aminoethanol are competitive inhibitors. L-serine dehydratase activity is strongly inhibited by o-phenanthraline. Pyridoxal phosphate is required for maximum activity.

Increase of L-serine dehydratase activity under gluconeogenic conditions in adult-rat hepatocytes cultured on collagen gel/nylon mesh.

Hormonal regulation of L-serine dehydratase [L-serine hydro-lyase (deaminating), EC 4.2.1.13] was studied in primary cultures of adult-rat hepatocytes. The hepatocytes were isolated by collagenase perfusion and maintained in culture on collagen-gel/nylon-mesh substrata. L-Serine dehydratase activity was measured with [14C]threonine as substrate. The enzyme activity in hepatocytes of normal adult rats was low and declined rapidly in culture in L-15 medium containing 0.1 micro M-insulin and even more in the presence of glucose. L-Serine dehydratase activity in hepatocytes of rats with streptozotocin-induced diabetes was initially 20-fold higher than that of normal rats, but fell rapidly to a low value by 4 days in culture. Hormonal regulation of the enzyme activity was manifested by treatment of the cultured hepatocytes with insulin (0.1 micro M), glucagon (0.3 micro M), dexamethasone (10 micro M) and combinations of these hormones. Either glucagon or dexamethasone in the absence of insulin enhanced the activity of L-serine dehydratase, but failed to do so in the presence of insulin. Treatment with both hormones resulted in a 2-3-fold increase in enzyme activity in culture on days 3 and 4. Under conditions in which the enzyme activity was enhanced, glucose production by the cultured hepatocytes was concomitantly increased. Glucose production resulted in part from gluconeogenesis from pyruvate and not entirely from glycogenolysis. The gluconeogenic conditions of culture resulted in a decrease in cellular lipids in the cultured hepatocytes, as evidenced by ultrastructural studies.

Regulation of hepatic L-serine dehydratase and L-serine-pyruvate aminotransferase in the developing neonatal rat.

1. The activities of l-serine dehydratase and l-serine-pyruvate aminotransferase were determined in rat liver during foetal and neonatal development. 2. l-Serine-pyruvate aminotransferase activity begins to develop in late-foetal liver, increases rapidly at birth to a peak during suckling and then decreases at weaning to the adult value. 3. l-Serine dehydratase activity is very low prenatally, but increases rapidly after birth to a transient peak. After a second transient peak around the time weaning begins, activity gradually rises to the adult value. Both of these peaks have similar isoenzyme compositions. 4. In foetal liver both l-serine dehydratase and l-serine-pyruvate aminotransferase activities are increased after injection in utero of glucagon or dibutyryl cyclic AMP. Cycloheximide or actinomycin D inhibited the prenatal induction of both enzymes and actinomycin D blocked the natural increase of l-serine dehydratase immediately after birth. Glucose or insulin administration also blocked the perinatal increase of l-serine dehydratase. 5. After the first perinatal peak of l-serine dehydratase, activity is increased by cortisol and this is inhibited by actinomycin D. After the second postnatal peak, activity is increased by amino acids or cortisol and this is insensitive to actinomycin D inhibition. Glucose administration blocks the cortisol-stimulated increase in l-serine dehydratase and also partially lowers the second postnatal peak of activity. 6. The developmental patterns of the enzymes are discussed in relation to the pathways of gluconeogenesis from l-serine. The regulation of enzyme activity by hormonal and dietary factors is discussed with reference to the changes in stimuli that occur during neonatal development and to their possible mechanisms of action.

Bacterial origin of alkaline l-serine dehydratase in French beans

Two l-serine dehydratases with pH optima at pH 5·5 and 9·0, respectively, have been identified in homogenates of French bean seedlings. The l-serine dehydratase activity with optimum at pH 9·0 increases as the plant extract ages and this correlates with bacterial growth in the extract; appearance of the enzyme in plant homogenates was retarded by the antibacterial agent, chloromycetin. The main bacteria associated with the plant extracts were identified as Erwinia herbicola and Pseudomonas fluorescens. Examination of cell free extracts of these bacteria show that the Pseudomonas fluorescens contains an active l-serine dehydratase. Whole cells of the bacterium show optimum activity towards l-serine at pH 9·0. It is concluded that in French bean homogenates this l-serine dehydratase may be of bacterial origin; in contrast, the enzyme with optimum at pH 5·5 appears to be a plant enzyme.

Evidence for the presence of two forms of L-serine dehydratase in rat liver

Lserine dehydrates (L-serine hydrolyase (deaminating), EC 4.2.1.13) has been highly purified from rat liver after inducing the enzyme either by feeding the animals with a high-protein diet [l-4] or by fasting them under simultaneous administration of cortisone [4], As reported by Schmidinger and Kroger [5] L-serine dehydratase activity rapidly decreases in rat liver if the animals are fed with a protein-free diet. However, the enzyme level does not reach zero, even after giving the diet for 7 days. The enzyme activity increases again, as soon as the rats are fasted or put on a diet of casein hydrolysate. The present paper deals with the demonstration of two forms of L-serine dehydratase from rat liver. The enzymes are different from each other concerning their molecular weight and heat stability. Five female rats (Sprague-Dawley strain; Ivanovas, Kisslegg/Allglu; weighing 1 lo-130 g) were put on a protein-free diet (protein content less than 1%) for 7 days and thereafter starved for 38 hr in order to get induction of the enzyme. The animals were then killed and their livers removed. These were homogenized (Ultra-Turrax) for 40 set in 4 volumes of 0.14 M KC1 containing 10” M EDTA and 1 OW5 M PALP. The homogenates were centrifuged (105,000 X g; 40 min) and the supernatants were combined. This crude extract is defined as the “induced” enzyme preparation. If the rats, on the other hand, were not starved after getting the protein-free diet for 7 days thus no induction took place the supernatants mentioned above result in an extract called the “non-induced” enzyme preparation. The assay for L-serine dehydratase was carried out according to Schmidinger and Kroger [5], except for the incubation period which

L-serine dehydratase (deaminase) of psychrophiles and mesophiles from polar and temperate habitats.

Escherichia coli and Pseudomonas aeruginosa were grown at several temperatures ranging from 0° to 37 °C and assayed at each temperature for L-serine dehydratase activity. The relationship between enzymatic activity and growth temperature is quite different for the mesophile and psychrophile. In addition, the stimulatory effect of natural plant lecithin upon serine deamination was different for each organism, suggesting that there is either a difference in permeability or that this substance has a more direct role in enzymic activity. Also included are data confirming the results of other workers.