Measuring cysteine biosynthesis activity from serine in extracts from sorghum [Zea mays], corn [Sorghum bicolor] and grass weeds, and their metolachlor susceptibility
2002
Hirase, K. (USDA, Agricultural Research Service, Mississippi (USA)) | Molin, W.T.
In vitro assay procedures for measuring the activity of cysteine biosynthesis from serine (CBS), which is a coupled reaction catalyzed by serine acetyltransferase and cysteine synthase, were developed using crude extracts from sorghum shoots. Cysteine biosynthesis from serine activ-ity was dependent on acetyl-CoA concentrations (up to 1.5mmolL**-1), serine (at least up to 20mmolL**-1) and sulfide (up to 0.25mmolL**-1), respectively, and was proportional to the protein concentration in the reaction mixture below 0.4mgmL**-1. The reaction rate was 6.6 nmolminE-1 per mg of protein during the first 5 min, but increased to 45.6 nmolminE-1 per mg of protein between 30 and 45 min after reaction initiation. Sorghum had the highest CBS total activity (222.4nmolminE-1 per g of fresh weight), and large crabgrass had the lowest CBS total activity (4.7 nmolminE-1 per g of fresh weight) when CBS activity in shoots was extracted from sorghum, corn, johnsongrass, barnyardgrass, goosegrass, green foxtail and large crabgrass. Similar results were obtained for CBS specific activity (nmolminE-1 per mg of protein).There was no correlation between total CBS activity and susceptibility to metolachlor; however, when corn was excluded, a correlation of R**2 = 0.690 was found. Flurazole seed treatment (1.25g per kg of seed) conferred metolachlor resistance by sorghum, and enhanced total CBS activity and non-protein thiol content by 27 and 61%, respectively. The increase in thiol content presumably contributed to metolachlor tolerance in sorghum. From these results, the difference in CBS activity partially contributes to the selectivity to metolachlor among certain grass species, and to the safening action of flurazole by increasing thiol content.
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