Purification and Characterization of Cystine Lyase a from Broccoli Inflorescence

Ukai, Koji; Sekiya, Jiro

doi:10.1271/bbb.61.1890

Cited by 14 publications

(20 citation statements)

References 9 publications

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“…Native PAGE was performed as described in Ref. 16. Protein concentrations were estimated using bovine serum albumin as a reference (17).…”

Section: Methodsmentioning

confidence: 99%

“…The active diluted cystine lyase fraction was then applied to a Cibacron blue 3G (Sigma) column equilibrated in TP buffer, and proteins were eluted with a linear gradient from 0 to 1 M NaCl. Active fractions were pooled and subjected to preparative native PAGE with activity staining and elution as described previously (16).…”

Section: Methodsmentioning

confidence: 99%

“…Lyases-A cystine-lyase activity was isolated from broccoli using a combination of previously established procedures (16) and additional Cibacron blue affinity chromatography. Two polypeptides migrating around 43 kDa by SDS-PAGE correlated with cystine lyase activity after elution from the Cibacron blue 3GA column with a linear salt gradient (data not shown).…”

Section: Isolation Of Broccoli Cystinementioning

confidence: 99%

“…A, native PAGE of pooled fractions containing cystine lyase activity after Cibacron blue chromatography. Lane 1 was stained with Coomassie Blue, while lane 2 was subjected to cystine lyase activity staining (16). B, SDS-PAGE of proteins (visualized by Coomassie Blue) eluted from a preparative native PAGE gel fraction that co-migrated with cystine lyase activity staining as described above.…”

Section: Isolation Of Broccoli Cystinementioning

confidence: 99%

“…The purification of a plant cystine lyase is not novel (1,16,25); however, the identification of a plant-derived nucleic acid sequence that encodes a cystine lyase, CORI3, certainly is. The isolation of BOCL3 and its corresponding cDNA clone by classical biochemical methods proved essential for the identification of the true function of the protein encoded by CORI3.…”

Section: Isolation Of Broccoli Cystinementioning

confidence: 99%

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A New Member of Plant CS-lyases

Jones

Manabe

Awazuhara

et al. 2003

Journal of Biological Chemistry

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“…Native PAGE was performed as described in Ref. 16. Protein concentrations were estimated using bovine serum albumin as a reference (17).…”

Section: Methodsmentioning

confidence: 99%

Section: Methodsmentioning

confidence: 99%

Section: Isolation Of Broccoli Cystinementioning

confidence: 99%

Section: Isolation Of Broccoli Cystinementioning

confidence: 99%

Section: Isolation Of Broccoli Cystinementioning

confidence: 99%

See 3 more Smart Citations

A New Member of Plant CS-lyases

Jones

Manabe

Awazuhara

et al. 2003

Journal of Biological Chemistry

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Activity staining of glutathione peroxidase after electrophoresis on native and sodium dodecyl sulfate polyacrylamide gels

2002

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Glutathione peroxidase (GSH-Px), from commercial bovine erythrocytes or ammonium sulfate fractionations (30-45%, 45-60%, 60-75% and 75-90% saturations) of ginger rhizome, was detected on polyacrylamide gels after native polyacrylamide gel electrophoresis (PAGE) or sodium dodecyl sulfate (SDS)-PAGE. The gel was submerged in a 50 mM Tris-HCl buffer (pH 7.9) containing 13 mM glutathione and 0.004% hydrogen peroxide with gentle shaking for 10-20 min. The GSH-Px activity was stained with a solution containing 1.2 mM 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) and 1.6 mM phenazine methosulfate (PMS) for 10 min. The clear zone of GSH-Px activity on a purple background was found in both native and SDS-PAGE gels. This fast and sensitive method can be used in the process of enzyme purification and characterization of mammalian or plant cells.

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Characterisation of the l‐Cystine β‐Lyase PatB from Phaeobacter inhibens: An Enzyme Involved in the Biosynthesis of the Marine Antibiotic Tropodithietic Acid

et al. 2017

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The l-cystine β-lyase from Phaeobacter inhibens is involved in the biosynthesis of the sulfur-containing antibiotic tropodithietic acid. The recombinant enzyme was obtained by heterologous expression in Escherichia coli and biochemically characterised by unambiguous chemical identification of the products formed from the substrate l-cystine, investigation of the substrate spectrum, determination of the enzyme kinetics, sequence alignment with closely related homologues and site-directed mutagenesis to identify a highly conserved lysine residue that is critical for functionality. PatB from P. inhibens is a new member of the small group of characterised l-cystine β-lyases and the first example of an enzyme with such an activity that is required for the biosynthesis of an antibiotic. A comparison of PatB to previously reported enzymes with l-cystine β-lyase activity from bacteria and plants is given.

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Purification and Characterization of Cystine Lyase a from Broccoli Inflorescence

Cited by 14 publications

References 9 publications

A New Member of Plant CS-lyases

A New Member of Plant CS-lyases

Activity staining of glutathione peroxidase after electrophoresis on native and sodium dodecyl sulfate polyacrylamide gels

Characterisation of the l‐Cystine β‐Lyase PatB from Phaeobacter inhibens: An Enzyme Involved in the Biosynthesis of the Marine Antibiotic Tropodithietic Acid

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