TY  - BOOK
AU  - Steiner,F.X.
AU  - Downey,R.J.
TI  - Isoelectric focusing and two-dimensional analysis of purified nitrate reductase from Aspergillus nidulans
KW  - ASPERGILLUS NIDULANS
KW  - ELECTROPHORESIS, POLYACRYLAMIDE GEL
KW  - ISOELECTRIC FOCUSING
KW  - KINETICS
KW  - MOLECULAR WEIGHT
KW  - NITRATE REDUCTASES
N2  - The assimilatory NADPH-nitrate oxidoreductase (EC 1.6.6.3)from Aspergillus nidulans was purified by means of affinity chromatography and analyzed by agarose isoelectric focusing and two-dimensional electrophoresis. NADPH-nitrate reductase activity was not activated by oxidation with potassium ferricyanide and was irreversibly inhibited by acrylamide. Electrophoresis of nitrate reductase in 7 percent polyacrylamide gels resulted in rapid loss of enzyme activity. Isoelectric focusing of purified enzyme in agarose gels resulted in one homogeneous band that exhibited NADPH-nitrate reductase, NADPH-cytochrome c reductase and reduced methyl viologen-nitrate reductase activities, which corresponded to an isoelectric point of 6.12 ± 0.05 at 22°C. Two-dimensional electrophoresis of focused nitrate reductase on SDS-polyacrylanide gel slabs yielded a single subunit of 54000 molecular weight. Acid treatment of the enzyme and subsequent isoelectric focusing resulted in a protein with a strongly acidic isoelectric point and reduced methyl viologen-nitrate reductase activity. It released another protein with a strongly basic isoelectric point which was inactive. It is postulated that the overall association of fiavoprotein protomers with both heme and cytochrome b1 components confers a small net negative charge upon the native heteromultimer and accounts for its slightly acidic isoelectric point. © 1982
UR  - https://drive.google.com/file/d/149u9PKXKF81PGGF0Q0d9gef3WnqeEgDp/view?usp=drivesdk
ER  -