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. 2000 Apr;122(4):1311–1322. doi: 10.1104/pp.122.4.1311

Figure 5.

Figure 5

Reconstitution of recombinant CYP79E1 and CYP79E2 using radiolabeled Tyr as substrate. Spheroblasts were isolated from E. coli cells expressing different constructs of CYP79E1 and CYP79E2, followed by temperature-induced Triton X-114 phase partitioning and reconstitution with S. bicolor NADPH-Cyt P450 oxidoreductase. Lanes 1 to 6 contain 20 μg protein/reaction mixture. Lane 7 contains 10 μg protein/reaction mixture. Lane 1, CYP79E1na, no NADPH; lane 2, CYP79E1na, no NADPH-Cyt P450 oxidoreductase; lane 3, CYP79E1na; lane 4, CYP79E1Δ (1–52)2E1(10aa); lane 5, CYP79E2lacZ(24aa); lane 6, expression vector pSP19g10L; lane 7, S. bicolor CYP79A1Δ (1–33)17α(8aa). Reaction mixtures were analyzed by TLC and the products formed monitored and quantified using a phosphor imager. The position of authentic standards is indicated. Oxime, (E)- and (Z)-p-Hydroxyphenylacetaldoxime; nitrile, p-hydroxyphenylacetonitrile.