FIG. 3.

SDS-PAGE (12.5%) of fractions obtained during purification of native and recombinant 3-hydroxypropionyl-CoA dehydratase. Proteins were stained with Coomassie blue. (A) Enzyme fractions during purification of the native enzyme from M. sedula. Lane 1, cell extract of autotrophically grown cells (20 μg); lane 2, after heat precipitation (20 μg); lane 3, after dialysis overnight (20 μg); lane 4, after Q-Sepharose chromatography (10 μg); lane 5, after carboxymethylcellulose chromatography (10 μg); lane 6, after Cibacron blue chromatography (the marked band is 3-hydroxypropionyl-CoA dehydratase; protein amount not determined); lane 7, molecular mass standard proteins. (B) Heterologous expression of the 3-hydroxypropionyl-CoA dehydratase gene from M. sedula in E. coli Rosetta 2 (DE3). Lane 1, whole cells before induction; lane 2, whole cells after 3 h of induced growth; lane 3, cell extract after heat precipitation (20 μg); lane 4, purified recombinant 3-hydroxypropionyl-CoA dehydratase after Ni²⁺ affinity column (10 μg); lane 5, molecular mass standard proteins.