Isozymic pattern of lactate dehydrogenase in cases of bancroftian filariasis

1992 ◽  
Vol 66 (2) ◽  
pp. 142-146
Author(s):  
S. Misra ◽  
T. M. Mahopatra ◽  
S. Rathaur
Keyword(s):  
Lactate Dehydrogenase ◽  
Gel Electrophoresis ◽  
Healthy Subjects ◽  
Polyacrylamide Gel Electrophoresis ◽  
Combined Treatment ◽  
Gel Filtration ◽  
Bancroftian Filariasis ◽  
Ldh Activity ◽  
Isozymic Pattern ◽  
Endemic Normal

ABSTRACTIsozymic patterns of lactate dehydrogenase (E.C. 1.1.1.27) by polyacrylamide gel electrophoresis (PAGE) were observed m various categories of filariasis and controls, i.e. asymptomatic microfilaraemia and symptomatic amicrofilaraemia, endemic normal and non-endemic normal. Lactate dehydrogenase (LDH) activity was also observed amongst the above categories of patients. An increase in enzyme activity and change in the isozymic pattern was observed in the above categories of filaria infected serum. LDH activity doubled in asymptomatic microfilaraemia whereas in symptomatic amicrofilaraemia the increase in LDH activity was thirtyfold. The isozymic pattern of microfilaraemia cases showed the presence of three bands B4, A1B3, A2 B2, which are quite thick as compared to normal healthy subjects, whereas the patients with symptomatic amicrofilaraemia showed marked elevation of serum LDH-4 or A3B1. The LDH was partially purified by combined treatment of (NH4)2SO4 fractionation and gel filtration. The isozymic pattern of purified LDH showed a similar pattern.

Tissue specific isozymes of alanopine dehydrogenase in the channeled whelk Busycotypus canaliculatum

1982 ◽  
Vol 60 (7) ◽  
pp. 1568-1572 ◽  
Author(s):  
William C. Plaxton ◽  
Kenneth B. Storey
Keyword(s):  
Lactate Dehydrogenase ◽  
Dehydrogenase Activity ◽  
Gel Electrophoresis ◽  
Polyacrylamide Gel Electrophoresis ◽  
Physiological Functions ◽  
Ldh Activity ◽  
Muscle Tissues ◽  
Proboscis Muscles ◽  
Octopine Dehydrogenase ◽  
Muscular Tissues

The tissues of the channeled whelk Busycotypus canaliculatum displayed activities of three glycolytic dehydrogenases, alanopine dehydrogenase (ADH), octopine dehydrogenase (ODH), and lactate dehydrogenase (LDH). ADH and ODH were present in all seven tissues (hepatopancreas, gill, kidney, and mantle, ventricle, foot, and proboscis muscles) tested. ADH was the major cytosolic dehydrogenase activity in all tissues except proboscis, while ODH was present in high activities only in the four muscular tissues. Significant LDH activity occurred only in the two muscles which perform sustained work, ventricle, and proboscis.Tissue specific isozymes of ADH were identified. Three forms, specific for hepatopancreas, gill–kidney, and muscle tissues, were separable by isoelectrofocusing (pI's 5.69, 5.58, and 5.93, respectively), chromatofocusing, and polyacrylamide gel electrophoresis. The three isozymes differed kinetically showing significant differences in apparent Km's for L-alanine (e.g., 8.84 ± 0.03, 10.64 ± 0.40, and 13.12 ± 0.65 mM for hepatopancreas, ventricle, and gill, respectively) and even greater differences in apparent Km's for glycine (619 ± 55, 1412 ± 115, and 2542 ± 230 mM for the above three tissues, respectively). The ratios Km(gly)/Km(ala) averaged 70, 192, and 132 for the three isozymes, hepatopancreas, gill–kidney and muscle, respectively.The physiological functions of ADH isozymes in the whelk may be similar to those of the M and H isozymes of LDH in vertebrates or the muscle and brain specific isozymes of ODH in cephalopod molluscs.

scholarly journals Characterization of [3H]palmitate- and [3H]ethanolamine-labelled proteins in the multicellular parasitic trematode Schistosoma mansoni

1988 ◽  
Vol 254 (2) ◽  
pp. 419-426 ◽  
Author(s):  
P M Wiest ◽  
E J Tisdale ◽  
W L Roberts ◽  
T L Rosenberry ◽  
A A F Mahmoud ◽  
...  
Keyword(s):  
Schistosoma Mansoni ◽  
Molecular Mass ◽  
Gel Electrophoresis ◽  
Free Amino ◽  
Protein Modification ◽  
Polyacrylamide Gel Electrophoresis ◽  
Gel Filtration ◽  
Reductive Methylation ◽  
Amide Linkage

Biosynthetic labelling experiments with cercariae and schistosomula of the multicellular parasitic trematode Schistosoma mansoni were performed to determine whether [3H]palmitate or [3H]ethanolamine was incorporated into proteins. Parasites incorporated [3H]palmitate into numerous proteins, as judged by SDS/polyacrylamide-gel electrophoresis and fluorography. The radiolabel was resistant to extraction with chloroform, but sensitive to alkaline hydrolysis, indicating the presence of an ester bond. Further investigation of the major 22 kDa [3H]palmitate-labelled species showed that the label could be recovered in a Pronase fragment which bound detergent and had an apparent molecular mass of 1200 Da as determined by gel filtration on Sephadex LH-20. Schistosomula incubated with [3H]ethanolamine for up to 24 h incorporated this precursor into several proteins; labelled Pronase fragments recovered from the three most intensely labelled proteins were hydrophilic and had a molecular mass of approx. 200 Da. Furthermore, reductive methylation of such fragments showed that the [3H]ethanolamine bears a free amino group, indicating the lack of an amide linkage. We also evaluated the effect of phosphatidylinositol-specific phospholipase C from Staphylococcus aureus: [3H]palmitate-labelled proteins of schistosomula and surface-iodinated proteins were resistant to hydrolysis with this enzyme. In conclusion, [3H]palmitate and [3H]ethanolamine are incorporated into distinct proteins of cercariae and schistosomula which do not bear glycophospholipid anchors. The [3H]ethanolamine-labelled proteins represent a novel variety of protein modification.

scholarly journals Biochemical characterization including amino acid and carbohydrate compositions of canine and human brain Thy-1 antigen

1981 ◽  
Vol 197 (3) ◽  
pp. 629-636 ◽  
Author(s):  
J L McKenzie ◽  
A K Allen ◽  
J W Fabre
Keyword(s):  
Monoclonal Antibody ◽  
Amino Acid ◽  
Human Brain ◽  
Affinity Chromatography ◽  
Gel Electrophoresis ◽  
Polyacrylamide Gel Electrophoresis ◽  
Sodium Dodecyl ◽  
Gel Filtration ◽  
Antibody Affinity ◽  
Lentil Lectin

Human and canine brain Thy-1 antigens were solubilized in deoxycholate and antigen activity was followed both by conventional absorbed anti-brain xenosera of proven specificity and by mouse monoclonal antibodies to canine and human Thy-1. It is shown that greater than 80% of Thy-1 activity in the dog and man binds to lentil lectin, that the mobility on sodium dodecyl sulphate/polyacrylamide-gel electrophoresis of canine and human Thy-1 is identical with that of rat Thy-1 and that the Stokes radius in deoxycholate of canine and human brain Thy-1 is 3.0 nm and 3.25 nm respectively. Both lentil lectin affinity chromatography followed by gel-filtration chromatography on the one hand and monoclonal antibody affinity chromatography on the other gave high degrees of purification of the brain Thy-1 molecule in the dog and man, resulting in single bands staining for both protein and carbohydrate on sodium dodecyl sulphate/polyacrylamide-gel electrophoresis (except for a slight contaminant of higher molecular weight staining for protein but not carbohydrate with human Thy-1 purified by lentil lectin and gel-filtration chromatography). Analysis of canine and human brain Thy-1 purified by monoclonal antibody affinity chromatography with additional gel filtration through Sephadex G-200 showed that these molecules had respectively 38% and 36% carbohydrate. The amino acid and carbohydrate compositions were similar to those previously reported for Thy-1 of the rat and mouse, the main point of interest being the presence in canine and human brain Thy-1 of N-acetylgalactosamine, which has been reported in rat and mouse brain Thy-1 but not in Thy-1 from other tissues.

scholarly journals Analysis of the forms of acetylcholinesterase from adult mouse brain

1975 ◽  
Vol 147 (2) ◽  
pp. 205-214 ◽  
Author(s):  
E D Adamson ◽  
S E Ayers ◽  
Z A Deussen ◽  
C F Graham
Keyword(s):  
Enzyme Activity ◽  
Mouse Brain ◽  
Gel Electrophoresis ◽  
Polyacrylamide Gel Electrophoresis ◽  
Gel Filtration ◽  
Acetylcholinesterase Activity ◽  
Total Activity ◽  
Polyacrylamide Gel ◽  
Soluble Extract ◽  
Molecular Sizes

The solubilization of 80% of the acetylcholinesterase activity of mouse brain was performed by repeated 2h incubations of homogenates at 37 degrees C in an aqueous medium. Analysis of the soluble extract by gel filtration on Sephadex G-200 showed that up to 80% of the enzyme activity was eluted in a peak which was estimated to consist of molecules of about 74000mol.wt. This peak was called the monomer form of the enzyme. After 3 days at 4 degrees C, the soluble extract was re-analysed and was eluted from the column in four peaks of about 74000, 155000, 360000 and 720000 mol.wt. Since the total activity of the enzyme in these peaks was the same as that in the predominantly monomer elution profile of fresh enzyme, we concluded that the monomer had aggregated, possibly into dimers, tetramers and octomers. Extracts of the enzyme were analysed by polyacrylamide-gel electrophoresis and the resulting multiple bands of enzyme activity on gels were shown to separate according to their molecular sizes, that is by molecular sieving. All these forms had similar susceptibilities to the inhibitors eserine, tetra-isopropyl pyrophosphoramide and compound BW 284c51 [1,5-bis-(4-allyldimethylammoniumphenyl)pentan-3-one dibromide]. Thus the forms of the enzyme in mouse brain which can be detected by gel filtration and polyacrylamide-gel electrophoresis may all be related to a single low-molecular-weight form which aggregates during storage. This supports similar suggestions made for the enzyme in other locations.

Purification and properties of mercuric reductase from Yersinia enterocolitica 138A14

1993 ◽  
Vol 39 (2) ◽  
pp. 193-200 ◽  
Author(s):  
Mohamed Blaghen ◽  
Dominique J. M. Vidon ◽  
Mohamed Said El Kebbaj
Keyword(s):  
Molecular Weight ◽  
Yersinia Enterocolitica ◽  
Gel Electrophoresis ◽  
Polyacrylamide Gel Electrophoresis ◽  
Gel Filtration ◽  
Mercury Resistance ◽  
Thiol Compounds ◽  
Mercuric Reductase ◽  
Purification And Properties ◽  
Layer Chromatography

A mercuric ion-reducing flavoprotein was purified from Yersinia enterocolitica 138A14 using dye matrix affinity chromatography. The purified enzyme had a characteristic absorption spectrum similar to those of flavin compounds, and FAD was detected as a part of the purified enzyme by thin-layer chromatography. Freshly purified preparations of the enzyme showed a single band on SDS polyacrylamide gel electrophoresis with a molecular weight of 70 000. The isolated enzyme had a molecular weight of about 200 000 as determined by gel filtration and disc gel electrophoresis. These results suggest an apparently trimeric structure of the enzyme. Dithiothreitol treatment disrupted the trimer into a dimeric structure of 140 000. Along with ageing, as well as limited proteolytic digestion, the enzyme evolved to give a dimeric molecule of 105 000 composed of two identical subunits of 52 000. The combination of the purified enzyme with HgCl2, or unexpectedly with merthiolate, oxidised the NADPH, which was followed spectrophotometrically. The Km for HgCl2 was dependent on the concentration of exogenous thiol compounds. A comparison of physical properties as well as kinetic characteristics indicated that the enzyme from Y. enterocolitica 138A14 is similar to mercuric reductases isolated from other mercury-resistant bacteria.Key words: Yersinia enterocolitica, mercury resistance, mercuric reductase.

Photoactivated linkage of catechol O-methyltransferase and S-adenosyl-L-methionine

1984 ◽  
Vol 62 (10) ◽  
pp. 964-969 ◽  
Author(s):  
Peter H. Yu
Keyword(s):  
Sodium Dodecyl Sulfate ◽  
Ultraviolet Light ◽  
Paper Chromatography ◽  
Gel Electrophoresis ◽  
Polyacrylamide Gel Electrophoresis ◽  
Sodium Dodecyl ◽  
Gel Filtration ◽  
Polyacrylamide Gel ◽  
Comt Inhibitors ◽  
Isoelectric Focussing

The formation of a stably linked complex of tritiated S-adenosyl-L-methionine (AdoMet) and catechol O-methyltransferase (COMT) has been achieved by irradiating the enzyme and ligand in Tris–HCl buffer (pH 7.5) with ultraviolet light at 254 nm. The reaction is specific as shown by a number of criteria. COMT inhibitors such as S-adenosylhomocysteine can block this photoactivated linkage. The [3H]AdoMet–COMT adduct has been shown to be a homogeneous protein by Sephadex gel filtration, sodium dodecyl sulfate – polyacrylamide gel electrophoresis, and isoelectric focussing. After extensive proteolysis of the [3H]AdoMet–COMT adduct with pronase P, one major labelled product was released. This fragment could be separated by paper chromatography and was shown to be chromatographically identical to that released from the [3H]AdoMet – phenylethanolamine N-methyltransferase adduct.

Partial characterization of an abnormal lactate dehydrogenase isoenzyme, LDH-1ex, in serum from a patient with hepatocellular carcinoma.

1989 ◽  
Vol 35 (5) ◽  
pp. 844-848
Author(s):  
D L Kalpaxis ◽  
E E Giannoulaki
Keyword(s):  
Hepatocellular Carcinoma ◽  
Lactate Dehydrogenase ◽  
Gel Electrophoresis ◽  
Polyacrylamide Gel Electrophoresis ◽  
Sodium Dodecyl ◽  
Gel Chromatography ◽  
Heat Stable ◽  
Single Polypeptide ◽  
Polypeptide Chains

Abstract Serum from a patient with hepatocellular carcinoma contained an abnormal isoenzyme of lactate dehydrogenase (LDH; EC 1.1.1.27), LDH-1ex, that on electrophoresis on 10-g/L agarose gel migrated anodally to the LDH-1 band. This isoenzyme was partly purified by ultrafiltration and preparative electrophoresis. Gel chromatography and sodium dodecyl sulfate/polyacrylamide gel electrophoresis studies of the resulting LDH-1ex preparation suggested that this isoenzyme is probably a tetramer made up of four single polypeptide chains (monomers), each having a molecular mass of about 32,000 Da. LDH-1ex was heat stable and reacted more readily with 2-hydroxybutyrate than did the slower migrating LDH-4 and LDH-5 isoenzymes. LDH-1ex showed no activity when lactate was omitted from the substrate solution or replaced by ethanol.

A three step purification procedure for human liver ferritin

1980 ◽  
Vol 58 (6) ◽  
pp. 494-498 ◽  
Author(s):  
M. Pagé ◽  
J. Lagueux ◽  
C. Gauthier
Keyword(s):  
Affinity Chromatography ◽  
Gel Electrophoresis ◽  
Human Liver ◽  
Polyacrylamide Gel Electrophoresis ◽  
Gel Filtration ◽  
Purification Procedure ◽  
Human Liver Ferritin ◽  
Normal Human Liver ◽  
Normal Human ◽  
Liver Ferritin

We describe a method for the purification of normal human liver ferritin by ultrafiltration, gel filtration on Sephacryl S-300, and affinity chromatography on DEAE-Affi Gel Blue. The purity of the ferritin obtained was verified by immunoelectrophoresis, Ouchterlony immunodiffusion, polyacrylamide gel electrophoresis, and electrofocusing. This rapid method yields 32% of the original ferritin.

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