The effect of target site mutation for acetolactate synthase (ALS)-inhibitor resistance on ALS activity was evaluated in a sulfonylurea-resistant (R) biotype of prickly lettuce with a proline173to histidine substitution in Domain A of the ALS enzyme. I50values for ALS inhibition by several ALS-inhibitor herbicides were determined for R and susceptible (S) biotypes. Results from both a standard ALS assay and a chloroplast assay for ALS activity showed that the R biotype also was cross-resistant to representatives of the imidazolinone (imazethapyr) and triazolopyrimidine (flumetsulam) families, but was not cross-resistant to the pyrimidinyl oxybenzoate (4,6-dimethoxypyrimidin-2-y 1-oxy-2-benzoic acid) tested. TheKm(pyruvate) was similar for ALS extracted from the R and S biotypes, suggesting that mutation for resistance did not alter pyruvate binding on the enzyme. However, specific activity of ALS from the R biotype was 57% less than specific activity of ALS from the S biotype, suggesting that the resistance mutation may affect enzyme function, expression, or stability. ALS from the R biotype was less sensitive to inhibition by the branched chain amino acids, valine, leucine, and isoleucine, than ALS from the S biotype. Reduced sensitivity to feedback inhibition was correlated with 70, 40, and 9% higher concentrations of valine, leucine, and isoleucine, respectively, on a per seed basis in R vs. S seed.

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Biochemical and physiological effects of target site resistance to herbicides inhibiting acetolactate synthase (ALS) were evaluated using sulfonylurea-resistant (R) and -susceptible (S) near isonuclearLactuca sativa‘Bibb’ lines derived by backcrossing the resistance allele fromLactuca serriolaL. intoL. sativa.Sequence data suggest that resistance inL. sativais conferred by a single-point mutation that encodes a proline197to histidine substitution in Domain A of the ALS protein; this is the same substitution observed in RL. serriola. Kmapp(pyruvate) values for ALS isolated from R and SL. sativawere 7.3 and 11.1 mM, respectively, suggesting that the resistance allele did not alter the pyruvate binding domain on the ALS enzyme. Both R and S ALS had greater affinity for 2-oxobutyrate than for pyruvate at the second substrate site. Ratios of acetohydroxybutyrate: acetolactate produced by R ALS across a range of 2-oxobutyrate concentrations were similar to acetohydroxybutyrate: acetolactate ratios produced by S ALS. Specific activity of ALS from RL. sativawas 46% of the specific activity from SL. sativa, suggesting that the resistance allele has detrimental effects on enzyme function, expression, or stability. ALS activity from R plants was less sensitive to feedback inhibition by valine, leucine, and isoleucine than ALS from S plants. Valine, leucine, and isoleucine concentrations were about 1.5 times higher in R seed than in S seed on a per gram of seed basis, and concentrations of valine and leucine were 1.3 and 1.6 times higher, respectively, in R leaves than in S leaves. Findings suggest that the mutation for resistance results in altered regulation of branched-chain amino acid synthesis.