Part:BBa_K5443028:Experience

Some preliminary evidence for successful expression of at least the C3H-Sam5 enzyme in pMQ3A comes from the brown diffusible pigment produced by E.coli DH10B(pMQ3A) cultures expressing these genes (Fig. 1 - see plats in first column on left); we believe this is a melanin-like pigment made from 'accidental' hydroxylation of tyrosine, which yields L-DOPA, which polymerises. There is precedent for this brown pigment in studies of C3H-HpaBC here (BBa_K5443003) and elsewhere (BBa_K1124011); the E.coli HpaBC enzyme seems to be more active in making the pigment than the Saccharothrix Sam5 enzyme.

Figure 1. Patched clones of plasmid variants.
Each plate shows patched colonies transformed with variants of our created plasmids pMQ3B and pMQ3C. Some patched colonies show diffusion of a brown pigment into the surrounding agar, suggesting a side-reaction of the enzyme HpaBC that produces the brown pigment L-DOPA.