Trp157Arg and Thr591Lys show impaired carboxylase and epoxidase activities
Sample . | Epoxidase activity, pmol/h × 10-2, % . | . | Carboxylase activity, pmol/h × 10-2 . | Epoxidation-carboxylation ratio . |
|---|---|---|---|---|
| Wild type | 27.2 | 97 | 25.0 | 1.1 |
| Wild type | 28.1 | 100 | 24.7 | 1.1 |
| Asp31Asn | 29.1 | 104 | 26.2 | 1.1 |
| Asp31Asn | 29.1 | 104 | 27.0 | 1.1 |
| Trp157Arg | 2.2 | 8 | 1.8 | 1.2 |
| Trp157Arg | 2.7 | 10 | 1.8 | 1.5 |
| Thr591Lys | ND | 0 | 0.2 | — |
| Thr591Lys | ND | 0 | 0.2 | — |
| Asp31Asn/Thr591Lys | ND | 0 | 0.2 | — |
| Asp31Asn/Thr591Lys | ND | 0 | 0.2 | — |
| Mock | ND | 0 | 0.2 | — |
| Mock | ND | 0 | 0.2 | — |
Sample . | Epoxidase activity, pmol/h × 10-2, % . | . | Carboxylase activity, pmol/h × 10-2 . | Epoxidation-carboxylation ratio . |
|---|---|---|---|---|
| Wild type | 27.2 | 97 | 25.0 | 1.1 |
| Wild type | 28.1 | 100 | 24.7 | 1.1 |
| Asp31Asn | 29.1 | 104 | 26.2 | 1.1 |
| Asp31Asn | 29.1 | 104 | 27.0 | 1.1 |
| Trp157Arg | 2.2 | 8 | 1.8 | 1.2 |
| Trp157Arg | 2.7 | 10 | 1.8 | 1.5 |
| Thr591Lys | ND | 0 | 0.2 | — |
| Thr591Lys | ND | 0 | 0.2 | — |
| Asp31Asn/Thr591Lys | ND | 0 | 0.2 | — |
| Asp31Asn/Thr591Lys | ND | 0 | 0.2 | — |
| Mock | ND | 0 | 0.2 | — |
| Mock | ND | 0 | 0.2 | — |
Lysates containing equivalent amounts of wild-type or mutant carboxylase protein (as determined by a quantitative Western analysis) were assayed for conversion of KH2 to KO (epoxidase activity) or for incorporation of [14C]-CO2 into a Glu substrate (carboxylase activity). Epoxidase activity was quantitated by reference to a KO standard, and carboxylase activity was determined by using a specific activity of 50 cpm/pmol for [14C]-CO2.
ND indicates not detected; —, not applicable.