Table III.
LPAAT activity of the amylose-affinity column purified MBP-tagged proteins
SDS-PAGE analysis showed that MBP, MBP-1848, MBP-1752, MBP-2060, and MBP-PlsC were expressed at high levels as 43-, 67-, 69-, 96-, and 70-kD bands, respectively, in both membrane (or the 100,000g pellet) and soluble (or the 100,000g supernatant) fractions of TB1 cell lysates. Comparable expression profiles were also seen for each of the D-to-G variants of MBP-tagged proteins relative to the corresponding nonmutagenized proteins.
| Protein | LPAAT, Specific Activitya
|
|
|---|---|---|
| 16:0 CoA | 18:0 CoA | |
| nmol min−1 mg−1 protein | ||
| MBP | 0.13 (0.039) | 0.10 (0.044) |
| MBP-2060 | 1.9 (0.98) | 1.9 (0.30) |
| MBP-2060D76G | 3.2 (1.1) | 3.8 (0.74) |
| MBP-1752 | 4.5 (0.55) | 7.0 (0.80) |
| MBP-1752D66G | 4.3 (0.66) | 2.6 (0.22) |
| MBP-1848 | 640 (160) | 5.2 (0.50) |
| MBP-1848D69G | 3.1 (1.2) | 1.9 (0.76) |
| MBP-PlsC | 638 (68) | 348 (120) |
| MBP-PlsCD77G | 2.4 (0.40) | 1.8 (0.38) |
a
Each value represents the average of three independent measurements. The numbers in parentheses denote the sd.