scholarly journals Preferential over-expression of the class alpha rat Ya2 glutathione S-transferase subunit in livers bearing aflatoxin-induced pre-neoplastic nodules. Comparison of the primary structures of Ya1 and Ya2 with cloned class alpha glutathione S-transferase cDNA sequences

1990 ◽  
Vol 268 (2) ◽  
pp. 295-302 ◽  
Author(s):  
J D Hayes ◽  
L A Kerr ◽  
D J Harrison ◽  
A D Cronshaw ◽  
A G Ross ◽  
...  
Keyword(s):  
Amino Acid ◽  
Aflatoxin B1 ◽  
Acquired Resistance ◽  
Amino Acid Sequences ◽  
Glutathione S Transferase ◽  
Over Expression ◽  
Cdna Sequences ◽  
Normal Rat ◽  
Primary Structures ◽  
Rat Livers

Normal rat liver expresses Ya (Mr 25,500), Yc (Mr 27,500) and Yk (Mr 25,000) Class Alpha glutathione S-transferase (GST) subunits. The Ya-type subunit can be resolved into two separate polypeptides, designated Ya1 and Ya2, by reverse-phase h.p.l.c. In rat livers that possess aflatoxin B1-induced pre-neoplastic nodules, a marked increase is observed in the expression of Ya1, Ya2, Yc and Yk; of these subunits, Ya2 exhibited the greatest increase in concentration. The Ya1 and Ya2 subunits isolated from nodule-bearing livers were cleaved with CNBr, and the purified peptides were subjected to automated amino-acid-sequence analysis. Differences in the primary structures of the two Ya GST subunits were found at positions 31, 34, 107 and 117. These data demonstrate that Ya1 and Ya2 are distinct polypeptides and are the products of separate genes. The amino acid sequences obtained from Ya1 and Ya2 were compared with the cloned cDNAs pGTB 38 [Pickett, Telakowski-Hopkins, Ding, Argenbright & Lu (1984) J. Biol. Chem. 259, 4112-4115] and pGTR 261 [Lai, Li, Weiss, Reddy & Tu (1984) J. Biol. Chem. 259, 5182-5188], which encode rat Ya-type subunits. From these comparisons it appears probable that Ya1 represents the GST subunit encoded by pGTR 261, whereas Ya2 represents the subunit encoded by pGTB 38. It is likely that the over-expression of Ya1 and Ya2 in nodule-bearing livers is of major significance in the acquired resistance of nodules to aflatoxin B1, since previous work [Coles, Meyer, Ketterer, Stanton & Garner (1985) Carcinogenesis 6, 693-697] has shown that the Ya-type GST subunit has high activity towards aflatoxin B1 8,9-epoxide.

scholarly journals Nucleotide and derived amino acid sequences of the major porin of Comamonas acidovorans and comparison of porin primary structures.

1991 ◽  
Vol 173 (7) ◽  
pp. 2196-2205 ◽  
Author(s):  
S Gerbl-Rieger ◽  
J Peters ◽  
J Kellermann ◽  
F Lottspeich ◽  
W Baumeister
Keyword(s):  
Amino Acid ◽  
Amino Acid Sequences ◽  
Comamonas Acidovorans ◽  
Primary Structures

scholarly journals Human liver glutathione S-transferase ψ. Chemical characterization and secondary-structure comparison with other mammalian glutathione S-transferases

1987 ◽  
Vol 243 (1) ◽  
pp. 61-67 ◽  
Author(s):  
S V Singh ◽  
A Kurosky ◽  
Y C Awasthi
Keyword(s):  
Amino Acid ◽  
Secondary Structure ◽  
Human Liver ◽  
Chemical Characterization ◽  
Compositional Analysis ◽  
Amino Acid Sequences ◽  
Terminal Region ◽  
Glutathione S Transferase ◽  
Structure Comparison ◽  
Liver Glutathione

The isolation and chemical characterization of the anionic human liver glutathione S-transferase (GST) psi (pI 5.5) are described and compared with other GST isoenzymes reported for rat and human. Amino acid compositional analysis, substrate specificity and isoelectric focusing indicated that GST psi is a unique isoenzyme form of GST. Strikingly, however, amino acid sequence analysis of the N-terminal region indicated that GST psi was identical with GST mu in the first 23 amino acid residues reported. It is likely that these two enzyme forms are at least partially structurally related. In order to investigate further the genetic relationship of GST psi to other reported GST isoenzymes, secondary-structure analysis was performed. Despite substantial differences in the N-terminal-region amino acid sequences of some of the GST isoenzymes, the secondary structure of all the isoenzymes is highly conserved at their N-termini. The general uniformity of the secondary structure of this enzyme class at their N-termini strongly indicated that the observed diversity of these isoenzymes probably occurred as a result of a mechanism of gene duplication followed by divergence rather than a mechanism of convergent evolution.

scholarly journals Comparison of the mRNA sequences for Pi class glutathione transferases in different hamster species and the corresponding enzyme activities with anti-benzo[a]pyrene-7,8-dihydrodiol 9,10-epoxide

1996 ◽  
Vol 318 (2) ◽  
pp. 533-538 ◽  
Author(s):  
Stellan SWEDMARK ◽  
Bengt JERNSTRÖM ◽  
Dag JENSSEN
Keyword(s):  
Cell Lines ◽  
Chinese Hamster Ovary Cells ◽  
Chinese Hamster ◽  
Amino Acid Sequences ◽  
Cho Cell ◽  
Glutathione S Transferase ◽  
V79 Cells ◽  
Ovary Cells ◽  
Cdna Sequences ◽  
Chinese Hamster V79 Cells

Glutathione S-transferase (GST) of class Pi (GST Pi) is known to detoxify the mutagenic and carcinogenic (+)-anti-benzo[a]pyrene-7,8-dihydrodiol 9,10-epoxide [(+)-anti-BPDE] by conjugation with glutathione. Previously, we have shown that Chinese hamster V79 cells contain GST Pi, but seem to lack the capacity to conjugate (+)-anti-BPDE, although these cells do conjugate other substrates with GSH [Romert, Dock, Jenssen and Jernström (1989) Carcinogenesis 10, 1701–1707; Swedmark, Romert, Morgenstern and Jenssen (1992) Carcinogenesis 13, 1719–1723; Swedmark and Jenssen (1994) Gene 139, 251–256]. In the present study we have compared four cell lines derived from different hamster species with respect to GST cDNA sequences and capacity to conjugate (+)- or (-)-anti-BPDE. The cell lines were V79 and Chinese hamster ovary cells (CHO), Armenian hamster lung (AHL) cells and baby hamster kidney (BHK) cells. The sequencing revealed a complete homology between the V79 and CHO cDNA for GST Pi, whereas the corresponding amino acid sequences predicted from the corresponding AHL and BHK cDNAs differed by six and nine amino acids, respectively, from the predicted V79 sequence. None of these changes alone was found to influence the xenobiotic substrate-binding site. The cytosolic fractions from BHK and AHL cells were found to catalyse conjugation of (+)-anti-BPDE with GSH, whereas the corresponding activity in CHO cells was non-detectable. As shown previously, V79 cells were devoid of activity towards (+)-anti-BPDE. All the cell lines studied demonstrated appreciable GST activity towards 1-chloro-2,4-dinitrobenzene, but no activity with (-)-anti-BPDE. The latter result suggests that GST Pi is the sole or predominant GST in these cell lines. This was confirmed by HPLC analysis of purified enzymes obtained by affinity chromatography. However, when the catalytic activities of the pure enzymes were determined, all four different GST Pi enzymes were found to be highly capable of conjugating (+)-anti-BPDE with GSH. This observation indicates the existence of an intracellular factor that selectively inhibits conjugation of (+)-anti-BPDE, but not of 1-chloro-2,4-dinitrobenzene in the V79 and CHO cell lines. This new phenomenon seems to be specific for Chinese hamster, since both these cell lines originate from this species.

scholarly journals Plant and Bacterial Metaxin-like Proteins: Novel Proteins Related to Vertebrate Metaxins Involved in Uptake of Nascent Proteins into Mitochondria

2020 ◽  
Author(s):  
Kenneth W. Adolph
Keyword(s):  
Amino Acid ◽  
Protein Import ◽  
Protein Domains ◽  
Amino Acid Sequences ◽  
Similar Degree ◽  
Glutathione S Transferase ◽  
Phylogenetic Groups ◽  
Bacterial Phyla ◽  
Novel Proteins ◽  
Plant Groups

ABSTRACTThe metaxins were originally identified as vertebrate proteins of the outer mitochondrial membrane involved in protein import into mitochondria. Metaxin proteins have also been found in diverse invertebrate phyla. The present study is concerned with examining whether metaxin-like proteins occur in plants and bacteria. Metaxin-like proteins were revealed by their homology with human metaxins and the possession of characteristic GST_Metaxin protein domains. The results demonstrate that metaxin-like proteins exist in plants that include a wide variety of angiosperms, both eudicots and monocots, and other plant groups. Metaxin-like proteins can also be detected in bacteria, particularly in the Proteobacteria phylum, but also in different bacterial phyla. Phylogenetic analysis indicates that plant metaxin-like proteins, bacterial metaxin-like proteins, and vertebrate metaxins form distinct phylogenetic groups, but are related. Metaxin-like proteins, however, are only distantly related to GSTs (glutathione S-transferase proteins). A similar degree of homology is found in aligning the amino acid sequences of plant and bacterial metaxin-like proteins with human metaxins 1, 2, and 3 and other vertebrate metaxins. The amino acid identities range from about 22%-28% for each alignment. The presence of two conserved protein domains, GST_N_Metaxin and GST_C_Metaxin, in both plant and bacterial metaxin-like proteins provides evidence that these proteins are related to the vertebrate and invertebrate metaxins. The metaxin-like proteins have predicted secondary structures that are dominated by alpha-helical segments, like the vertebrate and invertebrate metaxins.

The Drosophila GMII gene encodes a Golgi alpha-mannosidase II

1999 ◽  
Vol 112 (19) ◽  
pp. 3319-3330 ◽  
Author(s):  
C. Rabouille ◽  
D.A. Kuntz ◽  
A. Lockyer ◽  
R. Watson ◽  
T. Signorelli ◽  
...  
Keyword(s):  
Electron Microscopy ◽  
Amino Acid ◽  
Amino Acid Sequences ◽  
Gene Encoding ◽  
Cdna Sequences ◽  
Mouse Cdna ◽  
Immuno Electron Microscopy ◽  
Drosophila Cells ◽  
Drosophila Embryos ◽  
Similarity Expression

In this paper we show the organisation of the Drosophila gene encoding a Golgi alpha-mannosidase II. We demonstrate that it encodes a functional homologue of the mouse Golgi alpha-mannosidase II. The Drosophila and mouse cDNA sequences translate into amino acid sequences which show 41% identity and 61% similarity. Expression of the Drosophila GMII sequence in CHOP cells produces an enzyme which has mannosidase activity and is inhibited by swainsonine and by CuSO(4.) In cultured Drosophila cells and in Drosophila embryos, antibodies raised against a C-terminal peptide localise this product mainly to the Golgi apparatus as identified by cryo-immuno electron microscopy studies and by antibodies raised against known mammalian Golgi proteins. We discuss these results in terms of the possible use of dGMII as a Drosophila Golgi marker.

scholarly journals Molecular cloning and heterologous expression of a cDNA encoding a mouse glutathione S-transferase Yc subunit possessing high catalytic activity for aflatoxin B1-8,9-epoxide

1992 ◽  
Vol 285 (1) ◽  
pp. 173-180 ◽  
Author(s):  
J D Hayes ◽  
D J Judah ◽  
G E Neal ◽  
T Nguyen
Keyword(s):  
Catalytic Activity ◽  
Amino Acid ◽  
Amino Acid Sequence ◽  
Rat Liver ◽  
Cdna Clone ◽  
Aflatoxin B1 ◽  
Mouse Liver ◽  
High Catalytic Activity ◽  
Glutathione S Transferase ◽  
E Coli

Resistance to the carcinogenic effects of aflatoxin B1 (AFB1) in the mouse is due to the constitutive expression of an Alpha-class glutathione S-transferase (GST), YcYc, with high detoxification activity towards AFB1-8,9-epoxide. A cDNA clone (pmusGST Yc) for a murine GST Yc polypeptide has been isolated. Sequencing has shown the cDNA insert of pmusGST Yc to be 922 bp in length, with an open reading frame of 663 bp that encodes a polypeptide of M(r) 25358. The primary structure of the murine GST Yc subunit predicted by pmusGST Yc is in complete agreement with the partial amino acid sequence of the aflatoxin-metabolizing mouse liver GST described previously [McLellan, Kerr, Cronshaw & Hayes (1991) Biochem. J. 276, 461-469]. A plasmid, termed pKK-musGST Yc, which permits the expression of the murine Yc subunit in Escherichia coli, has been constructed. The murine GST expressed in E. coli was purified and found to be catalytically active towards several GST substrates, including AFB1-8,9-epoxide. This enzyme was also found to possess electrophoretic and immunochemical properties closely similar to those of the GST Yc subunit from mouse liver. However, the GST synthesized in E. coli and the constitutive mouse liver Alpha-class GST exhibited small differences in their chromatographic behaviour during reverse-phase h.p.l.c. Automated Edman degradation revealed alanine to be the N-terminal amino acid in the GST Yc subunit expressed in E. coli, whereas the enzyme in mouse liver possesses a blocked N-terminus. Although sequencing showed that the purified Yc subunit from E. coli lacked the initiator methionine, the amino acid sequence obtained over the first eleven N-terminal residues agreed with that predicted from the cDNA clone, pmusGST Yc. Comparison of the deduced amino acid sequence of the mouse Yc polypeptide with the primary structures of the rat Alpha-class GST enzymes revealed that it is more closely related to the ethoxyquin-induced rat liver Yc2 subunit than to the constitutively expressed rat liver Yc1 subunit. The significance of the fact that both mouse Yc and rat Yc2 exhibit high catalytic activity towards AFB1-8,9-epoxide, whereas rat Yc1 possesses little activity towards this compound, is discussed in terms of structure/function.

Glutathione S-transferase localization in aflatoxin B1-treated rat livers

1990 ◽  
Vol 11 (6) ◽  
pp. 927-931 ◽  
Author(s):  
David J. Harrison ◽  
Linda May ◽  
John D. Hayes ◽  
Gordon E. Neal
Keyword(s):  
Aflatoxin B1 ◽  
Glutathione S Transferase ◽  
Rat Livers

scholarly journals Ethoxyquin-induced resistance to aflatoxin B1 in the rat is associated with the expression of a novel Alpha-class glutathione S-transferase subunit, Yc2, which possesses high catalytic activity for aflatoxin B1-8,9-epoxide

1991 ◽  
Vol 279 (2) ◽  
pp. 385-398 ◽  
Author(s):  
J D Hayes ◽  
D J Judah ◽  
L I McLellan ◽  
L A Kerr ◽  
S D Peacock ◽  
...  
Keyword(s):  
Aflatoxin B1 ◽  
Neonatal Rat ◽  
High Catalytic Activity ◽  
Amino Acid Sequencing ◽  
Glutathione S Transferase ◽  
Adult Rats ◽  
Purification Scheme ◽  
Glutathione S Transferases ◽  
A Subunit ◽  
Rat Livers

A purification scheme has been devised for two ethoxyquin-inducible Alpha-class glutathione S-transferases (GSTs) which possess at least 25-fold greater activity towards aflatoxin B1 (AFB1)-8,9-epoxide than that exhibited by the GSTs (i.e. F, L, B and AA) that have been described previously. These two enzymes are both heterodimers and both contain a subunit of Mr 25,800. This subunit has been isolated from both of the GST isoenzymes and, after cleavage with CNBr, it has been subjected to automated amino acid sequencing. The primary structure of the Mr 25,800 subunit revealed that it forms part of a subfamily of Alpha-class GSTs which possess closest identity (about 92%) with the Yc subunit of apparent Mr 27,500, which is encoded by the recombinant cDNA clone pGTB42 [Telakowski-Hopkins, Rodkey, Bennett, Lu & Pickett (1985) J. Biol. Chem. 260, 5820-5825]. As these two GSTs possess less than 70% sequence identity with the Ya1 and Ya2 subunits, both of Mr 25,500, the constitutively expressed Yc subunit of Mr 27,500 has been renamed Yc1 and the ethoxyquin-inducible GST of Mr 25,800 has been designated Yc2. Using this nomenclature, the two GSTs with high activity for AFB1-8,9-epoxide are Ya1Yc2 and Yc1Yc2. Although evidence suggests that induction of Yc2 is responsible for the high detoxification capacity of livers from ethoxyquin-treated rats for AFB1-8,9-epoxide, resistance towards AFB1 may be multifactorial in this instance as dietary ethoxyquin also induces the Ya1, Ya2 and Yc1 subunits about 2.2-, 10.9- and 2.7-fold respectively. Besides the induction of GST by ethoxyquin, activity towards AFB1-8,9-epoxide is also elevated in the livers of neonatal rats and in livers that contain preneoplastic nodules. Western blotting experiments show that Yc2 is not present in hepatic cytosol from adult rats fed on normal diets but is expressed in neonatal rat livers and in the livers of adult rats that contain preneoplastic nodules that have arisen as a consequence of consuming diets contaminated with AFB1.

scholarly journals Genomic organization of three novel toxins from the scorpion Buthus martensi Karsch that are active on potassium channels

2000 ◽  
Vol 346 (3) ◽  
pp. 805-809 ◽  
Author(s):  
Li DAI ◽  
Jing-Jiang WU ◽  
Yi-Hua GU ◽  
Zheng-Dao LAN ◽  
Min-Hua LING ◽  
...  
Keyword(s):  
Amino Acid ◽  
Genomic Dna ◽  
Amino Acid Sequences ◽  
Signal Peptides ◽  
Amino Acid Residues ◽  
Specific Primers ◽  
Cdna Sequences ◽  
C Terminus ◽  
Partial Cdna ◽  
Rapid Amplification

The cDNA and genomic DNA of three novel toxins from the scorpion Buthus martensi Karsch that are active on K+ channels, designated BmKTX (where KTX is kaliotoxin), BmTX1 and BmTX2, were cloned and sequenced. On the basis of their known amino acid sequences, gene-specific primers for 3ʹ and 5ʹ rapid amplification of cDNA ends (RACE) were designed and synthesized. By overlapping the two partial cDNA sequences obtained by 3ʹ and 5ʹ RACE, their full-length cDNA sequences were completed. BmKTX encodes a signal peptide of 22 amino acid residues and a mature toxin of 38 residues, whereas BmTX1 and BmTX2 encode signal peptides of 20 and 21 residues respectively and a mature toxin of 38 residues for each. Their cDNA-deduced amino acid sequences were totally consistent with those determined except that the C-terminus of BmKTX had an additional Gly residue, which was removed during post-translational processing and was indispensable for the amidation of its C-terminal Lys residue. In addition, the first deduced amino acid for both BmTX1 and BmTX2 is Gln instead of pyro-Glu in the reported toxins, which obviously also undergoes post-translational processing. The genomic DNA species of these three toxins were also amplified by PCR, then cloned and sequenced. They all consisted of two exons disrupted by a small single intron. All of these introns were inserted within the signal peptides at position -6 for BmKTX and at position -5 for both BmTX1 and BmTX2 upstream of the mature toxins, and consisted of 87, 87 and 80 bp respectively.

scholarly journals Octopus S-crystallins with endogenous glutathione S-transferase (GST) activity: sequence comparison and evolutionary relationships with authentic GST enzymes

1995 ◽  
Vol 309 (3) ◽  
pp. 793-800 ◽  
Author(s):  
S H Chiou ◽  
C W Yu ◽  
C W Lin ◽  
F M Pan ◽  
S F Lu ◽  
...  
Keyword(s):  
Amino Acid ◽  
Sequence Comparison ◽  
Phylogenetic Trees ◽  
Expression System ◽  
Amino Acid Sequences ◽  
Nucleotide Sequencing ◽  
Major Protein ◽  
Glutathione S Transferase ◽  
Mammalian Tissues ◽  
Gst Activity

S-Crystallin is a major protein present in the lenses of cephalopods (octopus and squid). To facilitate the cloning of this crystallin gene, cDNA was constructed from the poly(A)+ mRNA of octopus lenses, and amplified by PCR for nucleotide sequencing. Sequencing of 10 of 15 positive clones coding for this crystallin revealed three distinct S-crystallin isoforms with 61-64% identity in nucleotide sequences and 42-58% similarity in amino acid sequences when compared with homologous crystallins in squid lenses. These charge-isomeric crystallins also show between 26 and 33% amino acid sequence identity to four major classes of glutathione S-transferase (GST), a major detoxification enzyme present in most mammalian tissues. For further analysis, expression of one of the S-crystallin cDNAs was carried out in the bacterial expression system pQE-30, and the S-crystallin protein produced in Escherichia coli was purified to homogeneity to determine the enzymic properties. We found that the expressed octopus S-crystallin possessed much lower GST activity than the authentic GSTs from other tissues. Sequence comparison and construction of phylogenetic trees for S-crystallins from squid and octopus lenses and various classes of GSTs revealed that S-crystallins represent a multigene family which is structurally related to Alpha-class GSTs and probably derived from the ancestral GST by gene duplication and subsequent multiple mutational substitutions.

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