Cloning of two 5-aminolevulinic acid synthase isozymes HemA and HemO from Rhodopseudomonas palustris with favorable characteristics for 5-aminolevulinic acid production

5-Aminolevulinic acid (ALA) synthase (ALAS) HemA from non-sulfur photosynthetic bacteria has been used for the ALA bioproduction, whereas the isoenzyme HemT/HemO is less studied and not used for ALA production. Two ALAS-encoding genes, hemA and hemO from Rhodopseudomonas palustris were cloned, purified and characterized. The ALASs had very high specific activity, 3.6 and 2.7 U/mg, respectively, and strong affinity for one of its substrates, succinyl-CoA, K (m) with values of 11 and 4.4 mu M, respectively. HemO retained up to 60 % maximum activity within a broad range of concentrations of hemin, while HemA kept only 20 % at 10 mu M hemin. Escherichia coli overexpressing HemA or HemO produced 5.7 and 6.3 g ALA/l, respectively, in a 5 l bioreactor.