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Identification of rat liver glutathione S-transferase Yb subunits by partial N-terminal sequencing after electroblotting of proteins onto a polyvinylidene difluoride membrane from an analytical isoelectric focusing gel.
Electrophoresis. 1990 Jul; 11(7):589-93.E

Abstract

Rat liver glutathione S-transferases were partially purified using S-hexyl glutathione affinity chromatography, followed by native isoelectric focusing employing a pH 7-11 or pH 3-10 gradient. Proteins were excised and eluted from the gel for determination of subunit composition using sodium dodecyl sulfate-polyacrylamide gel electrophoresis. In separate experiments, isoelectric focusing gels were equilibrated with a sodium dodecyl sulfate-containing buffer at high pH, and proteins on the gel were electroblotted onto a polyvinylidene difluoride membrane, utilizing graphite plates as electrodes. The membrane-bound proteins were visualized by Coomassie Brilliant Blue staining. The protein bands were then excised from the membrane and inserted into a gas phase sequenator for direct sequencing. N-Terminal sequences thus determined were compared with published cDNA sequences. The isoelectric points (pIs) and positions on the isoelectric focusing gel of Yb1Yb1, Yb1Yb2 and Yb2Yb2 subunits were determined. We have also located on the pH 3-10 focusing gel an N-terminal blocked glutathione S-transferase which has a molecular weight similar to Yb subunits.

Authors+Show Affiliations

Chang LH
Institute of Molecular Biology, Academia Sinica, Taipei, Taiwan, Republic of China.
Hsieh JC
No affiliation info available
Chen WL
No affiliation info available
Tam MF
No affiliation info available

MeSH

Amino Acid SequenceAnimalsBuffersChromatography, AffinityGlutathione TransferaseHydrolysisIsoelectric FocusingLiverMaleMolecular Sequence DataPolyvinylsRatsRats, Inbred Strains

Pub Type(s)

Comparative Study
Journal Article

Language

eng

PubMed ID

2226415

Citation

Chang, L H., et al. "Identification of Rat Liver Glutathione S-transferase Yb Subunits By Partial N-terminal Sequencing After Electroblotting of Proteins Onto a Polyvinylidene Difluoride Membrane From an Analytical Isoelectric Focusing Gel." Electrophoresis, vol. 11, no. 7, 1990, pp. 589-93.
Chang LH, Hsieh JC, Chen WL, et al. Identification of rat liver glutathione S-transferase Yb subunits by partial N-terminal sequencing after electroblotting of proteins onto a polyvinylidene difluoride membrane from an analytical isoelectric focusing gel. Electrophoresis. 1990;11(7):589-93.
Chang, L. H., Hsieh, J. C., Chen, W. L., & Tam, M. F. (1990). Identification of rat liver glutathione S-transferase Yb subunits by partial N-terminal sequencing after electroblotting of proteins onto a polyvinylidene difluoride membrane from an analytical isoelectric focusing gel. Electrophoresis, 11(7), 589-93.
Chang LH, et al. Identification of Rat Liver Glutathione S-transferase Yb Subunits By Partial N-terminal Sequencing After Electroblotting of Proteins Onto a Polyvinylidene Difluoride Membrane From an Analytical Isoelectric Focusing Gel. Electrophoresis. 1990;11(7):589-93. PubMed PMID: 2226415.
* Article titles in AMA citation format should be in sentence-case
TY - JOUR T1 - Identification of rat liver glutathione S-transferase Yb subunits by partial N-terminal sequencing after electroblotting of proteins onto a polyvinylidene difluoride membrane from an analytical isoelectric focusing gel. AU - Chang,L H, AU - Hsieh,J C, AU - Chen,W L, AU - Tam,M F, PY - 1990/7/1/pubmed PY - 1990/7/1/medline PY - 1990/7/1/entrez SP - 589 EP - 93 JF - Electrophoresis JO - Electrophoresis VL - 11 IS - 7 N2 - Rat liver glutathione S-transferases were partially purified using S-hexyl glutathione affinity chromatography, followed by native isoelectric focusing employing a pH 7-11 or pH 3-10 gradient. Proteins were excised and eluted from the gel for determination of subunit composition using sodium dodecyl sulfate-polyacrylamide gel electrophoresis. In separate experiments, isoelectric focusing gels were equilibrated with a sodium dodecyl sulfate-containing buffer at high pH, and proteins on the gel were electroblotted onto a polyvinylidene difluoride membrane, utilizing graphite plates as electrodes. The membrane-bound proteins were visualized by Coomassie Brilliant Blue staining. The protein bands were then excised from the membrane and inserted into a gas phase sequenator for direct sequencing. N-Terminal sequences thus determined were compared with published cDNA sequences. The isoelectric points (pIs) and positions on the isoelectric focusing gel of Yb1Yb1, Yb1Yb2 and Yb2Yb2 subunits were determined. We have also located on the pH 3-10 focusing gel an N-terminal blocked glutathione S-transferase which has a molecular weight similar to Yb subunits. SN - 0173-0835 UR - https://www.unboundmedicine.com/medline/citation/2226415/Identification_of_rat_liver_glutathione_S_transferase_Yb_subunits_by_partial_N_terminal_sequencing_after_electroblotting_of_proteins_onto_a_polyvinylidene_difluoride_membrane_from_an_analytical_isoelectric_focusing_gel_ DB - PRIME DP - Unbound Medicine ER -