Purification and characterization of three forms of glutathione transferase from Proteus mirabilis

Three forms of glutathione transferase (GST) with pI values of 6.0, 6.4 and 7.3 were isolated from Proteus mirabilis AF 2924 by glutathione-affinity chromatography followed by isoelectric focusing, and their structural, kinetic and immunological properties were investigated. Upon SDS/polyacrylamide-slab-gel electrophoresis, all forms proved to be composed of two subunits of identical (22,500) Mr. GST-6.0 and GST-6.4 together account for about 95% of the total activity, whereas GST-7.3 is present only in trace amounts. Extensive similarities have been found between GST-6.0 and GST-6.4. These include subunit molecular mass, amino acid composition, substrate specificities and immunological characteristics. GST-7.3 also cross-reacted (non-identity) with antisera raised against bacterial GST-6.0. None of the antisera raised against a number of human, rat and mouse GSTs cross-reacted with the bacterial enzymes, indicating major structural differences between them and the mammalian GSTs. This conclusion is further supported by c.d. spectra.

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Purification and characterization of a labile rat glutathione transferase of the Mu class

Biochemical Journal ◽

10.1042/bj2600789 ◽

1989 ◽

Vol 260 (3) ◽

pp. 789-793 ◽

Cited By ~ 24

Author(s):

A Kispert ◽

D J Meyer ◽

E Lalor ◽

B Coles ◽

B Ketterer

Keyword(s):

Amino Acid ◽

Gel Electrophoresis ◽

Glutathione Transferase ◽

Polyacrylamide Gel Electrophoresis ◽

Amino Acid Residues ◽

Purification And Characterization ◽

Terminal Amino Acid ◽

Terminal Amino ◽

Cnbr Cleavage

A labile GSH transferase homodimer termed 11-11 was purified from rat testis by GSH-agarose affinity chromatography followed by anion-exchange f.p.l.c. The enzyme is unstable in the absence of thiol(s) and has relatively low affinity for both 1-chloro-2,4-dinitrobenzene (Km 4.4 mM) and GSH (Km(app.) 4.4mM). Its mobility on SDS/polyacrylamide-gel electrophoresis is slightly less than that of subunits 3 and 4 and its pI is 5.2. Subunit 11 has a blocked N-terminal amino acid residue, but after CNBr cleavage fragments accounting for 113 amino acid residues were sequenced and showed 65% homology with corresponding sequences in subunit 4, indicating that it is a member of the Mu family. GSH transferase 11 is a major isoenzyme in testis, epididymis, prostate and brain and present at lower concentrations in other tissues.

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Purification and characterization of the major glutathione transferase from adult toad (Bufo bufo) liver

Biochemical Journal ◽

10.1042/bj2890417 ◽

1993 ◽

Vol 289 (2) ◽

pp. 417-422 ◽

Cited By ~ 16

Author(s):

A Aceto ◽

B Dragani ◽

T Bucciarelli ◽

P Sacchetta ◽

F Martini ◽

...

Keyword(s):

Amino Acid ◽

Glutathione Transferase ◽

Total Activity ◽

Bufo Bufo ◽

Terminal Sequence ◽

Purification And Characterization ◽

Terminal Amino ◽

Toad Bufo ◽

Toad Bufo Bufo

Five forms of glutathione transferase (GST) were resolved from the cytosol of adult common toad (Bufo bufo) liver by GSH-affinity chromatography followed by isoelectric focusing. The major enzyme (GST-7.64; 55% of total activity bound to the column) has a pI value of 7.64, is composed of two subunits each with a molecular mass of 23 kDa, and has the N-terminal amino acid residue blocked. GST-7.64 has also been characterized with respect to amino acid composition, substrate specificity, inhibition characteristics, c.d. spectra and immunological reactivity. The N-terminal sequence of some peptides obtained after tryptic digestion has also been determined. All together the results obtained suggest that the major toad liver GST is distinct from any known GST, including microbial, plant and mammalian GSTs.

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Expression, purification, and characterization of a membrane-bound d-amino acid dehydrogenase from Proteus mirabilis JN458

Biotechnology Letters ◽

10.1007/s10529-017-2388-0 ◽

2017 ◽

Vol 39 (10) ◽

pp. 1559-1566 ◽

Cited By ~ 2

Author(s):

Jinjin Xu ◽

Yajun Bai ◽

Taiping Fan ◽

Xiaohui Zheng ◽

Yujie Cai

Keyword(s):

Amino Acid ◽

Proteus Mirabilis ◽

Purification And Characterization ◽

Acid Dehydrogenase ◽

Amino Acid Dehydrogenase ◽

Membrane Bound

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Purification and characterization of an exoglucanase from Streptomyces flavogriseus

Canadian Journal of Microbiology ◽

10.1139/m84-183 ◽

1984 ◽

Vol 30 (9) ◽

pp. 1171-1178 ◽

Cited By ~ 42

Author(s):

C. R. Mackenzie ◽

D. Bilous ◽

K. G. Johnson

Keyword(s):

Isoelectric Focusing ◽

Gel Electrophoresis ◽

Extracellular Enzymes ◽

Main Product ◽

Polyacrylamide Gel Electrophoresis ◽

Sodium Dodecyl ◽

Gel Permeation Chromatography ◽

Purification And Characterization ◽

Gel Permeation

Streptomyces flavogriseus, a mesophilic actinomycete, produces high levels of extracellular enzymes capable of hydrolyzing cellulose and xylan. One such enzyme, an exoglucanase, has been purified to molecular homogeneity by a sequence involving DEAE Bio-Gel A chromatography, gel permeation chromatography on Bio-Gel P-60, preparative isoelectric focusing, and concanavalin A affinity chromatography. This purification sequence disclosed the presence of several distinct endoglucanase and xylanase fractions. Homogeneity of the purified enzyme was demonstrated by analytical isoelectric focusing and sodium dodecyl sulphate – polyacrylamide gel electrophoresis. The purified enzyme had a molecular weight of approximately 45 000 and an isoelectric point of 4.15. The enzyme demonstrated negligible activity with carboxymethylcellulose as the substrate. It was able to extensively hydrolyse acid-swollen cellulose; the main product of enzyme action was cellobiose.

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Prothrombin Metz : Purification and Characterization of a Variant of Human Prothrombin

10.1055/s-0038-1665670 ◽

1979 ◽

Author(s):

M Ribieto ◽

J Elion ◽

D Labie ◽

F Josso

Keyword(s):

Molecular Weight ◽

Gel Electrophoresis ◽

Polyacrylamide Gel Electrophoresis ◽

Factor Xa ◽

Polyacrylamide Gel ◽

Cleavage Pattern ◽

Purification And Characterization ◽

Double Immunodiffusion ◽

The Family

For the purification of the abnormal prothrombin (Pt Metz), advantage has been taken of the existence in the family of three siblings who, being double heterozygotes for Pt Metz and a hypoprothrombinemia, have no normal Pt. Purification procedures included barium citrate adsorption and chromatography on DEAE Sephadex as for normal Pt. As opposed to some other variants (Pt Barcelona and Madrid), Pt Metz elutes as a single symetrical peak. By SDS polyacrylamide gel electrophoresis, this material is homogeneous and appears to have the same molecular weight as normal Pt. Comigration of normal and abnormal Pt in the absence of SDS, shows a double band suggesting an abnormal charge for the variant. Pt Metz exhibits an identity reaction with the control by double immunodiffusion. Upon activation by factor Xa, Pt Metz can generate amydolytic activity on Bz-Phe-Val-Arg-pNa (S2160), but only a very low clotting activity. Clear abnormalities are observed in the cleavage pattern of Pt Metz when monitored by SDS gel electrophoresis. The main feature are the accumulation of prethrombin l (Pl) and the appearance of abnormal intermediates migrating faster than Pl.

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