In Kook Park

Characterization of antimicrobial agents extracted from Asterina pectinifera

The extracts from the starfish Asterina pectinifera obtained using various organic solvents and distilled water were tested for their antimicrobial activities against fungi and yeasts. The methanol and water extracts were found to be the most active and Aspergillus spp. and Cryptococcus neoformans proved to be sensitive species. The extracts are polar, stable to a wide range of pH and a high temperature, and possess a very distinctive UV spectrum. The colour reactions for functional groups of antimicrobial substances indicate the presence of some distinctive chemical properties common to steroidal oligoglycosides which results suggest that the antimicrobial activity is due to structures common to steroidal oligoglycosides.

Effect of prolonged starvation on the activities of malic enzyme and acetylcholinesterase in tissues of Japanese quail

During starvation muscle protein degradation is increased but the mechanism for this is uncertain. In this study Japanese quail were starved for 5 days and the activities of malic enzyme and acetylcholinesterase were determined in various tissues. SDS-polyacrylamide gel electrophoresis showed that the soluble proteins with molecular weights corresponding to 160, 120, 108, 99 and 38 kDa were absent in the liver of the starved group. In the pectoral muscle the soluble proteins with molecular weights corresponding to 69, 41 and 34 kDa were missing. The activity of malic enzyme in the liver, heart and pectoral muscle of the starved group decreased markedly whereas that of acetylcholinesterase increased markedly in the pectoral muscle (P < 0.005). It is concluded that in prolonged starvation acetylcholinesterase synthesis may be induced in tissues being subjected to protein catabolism and that this enzyme may be involved as a protease in protein degradation.

Effect of niacin deficiency on the thermal stability of NAD- and NADP-dependent dehydrogenases in liver and pectoral muscle of Japanese quail

The objective of this study was to determine whether the concentration of pyridine nucleotides in muscle and liver tissue of quail affected the heat stability of aldolase and selected enzymes involved in the oxidation-reduction of these cofactors. The thermal stability of malic enzyme, glyceraldehyde-3-phosphate dehydrogenase, lactic dehydrogenase, and aldolase in liver, and in pectoral muscle of quail was studied at incubation temperatures ranging from 27 to 60 degrees C. The concentrations of liver NAD, NADP, NADPH and the thermal inactivation of liver malic enzyme, glyceraldehyde-3-phosphate dehydrogenase, lactic dehydrogenase, and aldolase were not affected by niacin deficiency. In contrast, pectoral muscle glyceraldehyde-3-phosphate dehydrogenase in the niacin deficient quail compared to that of the controls had a markedly reduced thermal stability. This was associated with a corresponding decrease in the concentration of NAD and possibly NADPH. However, lactic dehydrogenase and aldolase activities were not affected. A similar pattern of heat inactivation was obtained when dialysed muscle and liver extracts were spiked with NAD or NADP. In these studies, NAD(P) protected muscle glyceraldehyde-3-phosphate dehydrogenase against heat inactivation to a much greater degree than that obtained with the other enzymes from muscle or liver tissue. These results suggest a causative relationship between the thermal stability of glyceraldehyde-3-phosphate dehydrogenase and coenzyme status in pectoral muscle tissue. This effect of niacin deficiency on the thermal stability of enzymes appears to be quite selective and specific.

Effects of 6-aminonicotinamide on levels of soluble proteins and enzyme activities in various tissues of Japanese quail

The effects of 6-aminonicotinamide (6-AN) on the levels of soluble proteins and enzyme activities in various tissues of Japanese quail were investigated. SDS-polyacrylamide gel electrophoresis showed that the soluble proteins with molecular masses corresponding to 160.4 and 52.5 kDa were either missing or present at lower concentrations in the brain of the 6-AN treated group compared to those in the control group. The soluble liver proteins with molecular masses 200, 120 and 70.5 kDa were missing in the treated group compared to those in the control while those of a molecular mass 15.1 kDa were found to be present at higher concentrations. Similarly, treatment with 6-AN decreased the concentration of soluble proteins in pectoral muscle with molecular masses 92.3, 54.5, 43.5, 41.2, 34.5, 27.5, 20.1 and 17.5 kDa and increased those with molecular masses 96.5, 37.7, 25.0, 19.3, 16.6, 13.8 and 10.8 kDa. In the heart, soluble proteins with molecular mass 84.6 kDa were increased. There was a marked reduction in the treatment group in the concentration of NAD in pectoral muscle but not in other tissues. A similar observation was also made with total RNA levels. The specific activity of malic enzyme was markedly increased by 6-AN treatment in the kidney and pectoral muscle but reduced in the liver. 6-Phosphogluconate dehydrogenase and lactate dehydrogenase activities were markedly reduced in the liver. Glyceraldehyde-3-phosphate dehydrogenase activity was significantly decreased in liver and pectoral muscle. NAD glycohydrolase activity was markedly decrease in pectoral muscle. Acetylcholinesterase activity was markedly reduced in liver but was enhanced in pectoral muscle. The results suggest that the metabolic actions of 6-AN are specific for certain proteins in the liver and muscle with the effect being most pronounced in muscle. The effects are also quite distinct from those by its analogue 3-acetylpyridine.

Effects of the neurotoxin 3-acetylpyridine on levels of soluble proteins and enzyme activities in various tissues of Japanese quail

(1) The effects of the neurotoxin 3-acetylpyridine on the levels of soluble proteins and enzyme activities in various tissues of Japanese quail were investigated. (2) SDS-polyacrylamide gel electrophoresis showed that in the brain the soluble proteins with a molecular mass corresponding to 18 kDa were increased in quail treated with this toxin. The soluble liver proteins with the largest molecular masses (200, 120, 98, 80.5 and 58 kDa) were either missing or present at lower concentrations in the treated group compared to those in the controls while those of lower molecular mass (62, 55, 45, 36.5 and 24 kDa) were found to be present in higher concentrations. Similarly, treatment with 3-acetylpyridine tended to decrease the concentration of soluble proteins in pectoral muscle having a high molecular mass (160, 98, 60, 33, 30.5, 22 and 14 kDa) and to increase those having a low molecular mass (26, 20, 19.5 and 16 kDa). (3) There was a marked reduction in the treatment group in the concentration of NAD in pectoral muscle but not in other tissues. A similar observation was also made with total RNAs levels. (4) The specific activity of malic enzyme and glyceraldehyde-3-phosphate dehydrogenase was markedly reduced in the liver and pectoral muscle of the treatment group but was not affected in other tissues. The specific activity of 6-phosphogluconate dehydrogenase was significantly lower in the liver only, and that of lactic dehydrogenase and acetylcholinesterase was not affected in any of the tissues examined. (5) The results suggest that the metabolic actions of 3-acetylpyridine are quite distinct from those shown by niacin deficiency and its analog such as 6-aminonicotinamide.

Effects of niacin deficiency on the levels of soluble proteins and enzyme activities in various tissues of Japanese quail.

The effects of niacin deficiency on the levels of soluble proteins and enzyme activities of Japanese quail have been investigated. SDS-polyacrylamide gel electrophoresis revealed that in the pectoral muscle the soluble proteins with molecular masses corresponding to 181, 128, 93, 76, 72, 62, 56, 43, 41, 28 and 20 kDa were present in lower amounts but those of 60, 50 and 37 kDa were present in higher amounts. In the heart the soluble proteins with a molecular mass of 181 kDa were present in lower amounts and in the brain those of 43 kDa were present in lower amounts but those of 221 kDa were present in higher amounts. In the intestine the soluble proteins with molecular masses corresponding to 181, 102, 83, 74, 72, 44 and 40 kDa were present in lower amounts but those of 41 kDa and 18 kDa were present in higher amounts. There was a marked reduction in the level of NAD and NADPH in the pectoral muscle of niacin deficient quail but not in other tissues. The specific activity of glyceraldehyde-3-phosphate dehydrogenase decreased markedly both in the liver and pectoral muscle of niacin deficient quail whereas that of 6-phosphogluconate dehydrogenase and malic enzyme decreased markedly in the liver or pectoral muscle, respectively. In contrast, the specific activity of acetylcholinesterase and carboxypeptidase increased markedly in the liver or the pectoral muscle, respectively. The results suggest that a severe niacin deficiency exerted specific effects on levels of some soluble proteins particularly in the pectoral muscle and intestine and on activities of certain enzymes in the liver and the pectoral muscle.

The coenzyme thiamine pyrophosphate inhibits the self-splicing of the group I intron

Effects of the coenzyme thiamine pyrophosphate and its analogs on the inhibition of self-splicing of primary transcripts of the phage T4 thymidylate synthase gene (td) were investigated. Of all compounds tested, the coenzyme thiamine pyrophosphate was the most potent inhibitor and the order of inhibitory efficiency for compounds tested was as follows: thiamine pyrophosphate>thiamine monophosphate>thiamine>thiochrome. Increasing guanosine concentration overcame the suppression of self-splicing by thiamine pyrophosphate close to the level of normal splicing. Kinetic analysis demonstrated that thiamine pyrophosphate acts as a competitive inhibitor for the td intron RNA with a Ki of 2.2mM. The splicing specificity inhibition by thiamine pyrophosphate is predominantly due to changes in Km.

Pyridoxal phosphate inhibits the group I intron splicing

The coenzyme pyridoxal phosphate and its analogs were tested for inhibition of the self-splicing of primary transcripts of the phage T4 thymidylate synthase gene (td). Of all compounds examined, the pyridoxal phosphate was the most potent inhibitor and the order of inhibitory efficiency for compounds tested was as follows: pyridoxal phosphate > pyridoxal > pyridoxine > pyridoxamine > pyridoxic acid. Increasing Mg2+ concentration up to 14 mM overcame the suppression of self-splicing by pyridoxal phosphate up to 95% of the level of normal splicing, implying its interference with effective catalysis of Mg2+. The kinetic analysis demonstrated that pyridoxal phosphate acts as a mixed type noncompetitive inhibitor for the td intron RNA with a Ki of 11.8 mM. The specificity of the splicing inhibition by pyridoxal phosphate is predominantly due to increases in Km and decreases in Vmax values. (Mol Cell Biochem xxx: 17–34, 2005)

Purification of soluble acetylcholinesterase from Japanese quail brain by affinity chromatography.

The purification of a soluble acetylcholinesterase from Japanese quail brain using affinity chromatography on concanavalin A-Sepharose and edrophonium-Sepharose is described. The affinity matrix was synthesized by coupling an inhibitor edrophonium to epoxy-activated Sepharose. Acetylcholinesterase was purified 10,416-fold with a specific activity of 2500 U/mg protein. Polyacrylamide gel electrophoresis in the presence of sodium dodecyl sulfate and mercaptoethanol gave only one band with a molecular weight of 62.5 kDa. The molecular weight of the purified acetylcholinesterase was estimated to be 245.5 kDa by gel chromatography on Sephacryl S-200 under nondenaturing conditions. Based on the molecular weight obtained by both SDS-PAGE and gel filtration the purified acetylcholinesterase was assumed to be a tetrameric form.

Effects of nicotinamide coenzymes on the stability of enzyme activities and proteins in niacin-deficient quail tissues against trypsin treatment.

The stability of liver and muscle enzymes and proteins in niacin-deficient quail towards trypsin treatment in the presence and absence of coenzymes, NAD or NADP, was characterized. The protection of liver dehydrogenases by coenzymes was low when they are subjected to trypsin digestion for 60 min. In contrast, in the muscle there was substantial protection against trypsin inactivation of glyceraldehyde-3-phosphate dehydrogenase by NAD and of 6-phosphogluconate dehydrogenase by NADP. Among all enzymes tested, glyceraldehyde-3-phosphate dehydrogenase showed the greatest protection against trypsin inactivation by NAD. SDS-polyacrylamide gel electrophoresis demonstrated that muscle proteins from the niacin-deficient group were more substantially protected compared to control and pair-fed groups when liver and muscle extracts were spiked with NAD and subjected to trypsin digestion. Overall results suggest that niacin deficiency exerted specific destabilizing effects on the stability of enzymes and proteins in muscle.