Properties of tryptophan indole-lyase from a piezophilic bacterium, Photobacterium profundum SS9

Tryptophan indole-lyase (Trpase), PBPRA2532, from Photobacterium profundum SS9, a piezophilic marine bacterium, has been cloned, expressed in Escherichia coli, and purified. The P. profundum Trpase (PpTrpase) exhibits similar substrate specificity as the enzyme from E. coli (EcTrpase). PpTrpase has an optimum temperature for activity at about 30 degrees C, compared with 53 degrees C for EcTrpase, and loses activity rapidly (t(1/2) similar to 30 min) when incubated at 50 degrees C, while EcTrpase is stable up to 65 degrees C. PpTrpase retains complete activity when incubated more than 3 h at 0 degrees C, while EcTrpase has only about 20% remaining activity. Under hydrostatic pressure, PpTrpase remains fully active up to 100 MPa (986 atm), while EcTrpase exhibits only about 10% activity at 100 MPa. PpTrpase forms external aldimine and quinonoid intermediates in stopped-flow experiments with L-Trp, S-Et-L-Cys, S-benzyl-L-Cys, oxindolyl-L-Ala, L-Ala and L-Met, similar to EcTrpase. However, with L-Trp a gem-diamine is observed that decays to a quinonoid complex. An aminoacrylate is observed with L-Trp in the presence of benzimidazole, as was seen previously with EcTrpase [28] but not with S-Et-L-Cys. The results show that PpTrpase is adapted for optimal activity in the low temperature, high pressure marine environment. (C) 2010 Elsevier Inc. All rights reserved.