ISHS Acta Horticulturae 650: I International Symposium on Saffron Biology and Biotechnology
EXPRESSION OF L-LACTATE DEHYDROGENASE ISOENZYMES DURING ROOT DEVELOPMENT IN CROCUS SATIVUS L. CORM |
| Authors: | J. Keyhani, E. Keyhani, N. Sattarahmady |
| Keywords: | cellular respiration, cytochrome c, electron transfer, flavocytochrome b2, isoenzymes, non-denaturing gel electrophoresis, potassium ferricyanide |
| Abstract:
Flavocytochrome b2 activity was detected in Crocus sativus L. corm extracts prepared before and at 10 and 30 days after rooting. The activity was measured spectrophotometrically by following the reduction of cytochrome c at 550 nm or that of potassium ferricyanide at 420 nm. pH activity profiles revealed optima at pH 5.5, 7.5 and 9.5 before rooting, at pH 5.5 and 9.0 after 10 days rooting, and at pH 5.5, 7.5 and 9.5 after 30 days rooting, using potassium ferricyanide as substrate. Results were similar when cytochrome c was used as substrate, with optima at slightly higher pHs (7.0, 8.0, 9.0-10.0). Kinetic parameters (Km, Vmax) were measured for each extract at pH optima. Results showed that the catalytic efficiency at a given pH would change as rooting was taking place. Using potassium ferricyanide as substrate, at pH 5.5 the catalytic efficiency (expressed per mg protein in the extract) was 2.1 ± 0.3 min-1 before rooting, 8.1 ± 0.8 min-1 after 10 days rooting, and 2.5 ± 0.4 min-1 after 30 days rooting. At pH 7.5, it was 5.7 ± 0.4 min-1 before rooting, and 9.2 ± 0.8 min-1 after 30 days rooting. At pH 9.5, it was 0.4 ± 0.04 min-1 before rooting, 2.1 ± 0.3 min-1 (pH 9.0) after 10 days rooting, and 6.0 ± 0.8 min-1 after 30 days rooting. When the extracts were submitted to non-denaturing gel electrophoresis, activity staining for lactate dehydrogenase revealed three distinct bands before rooting, two bands after 10 days rooting, and three bands after 30 days rooting. Data suggested the presence of isoenzymes of lactate dehydrogenase selectively expressed before and during rooting in C. sativus corms. This may be relevant to the metabolic control of the cell respiration under these conditions. | |
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