This paper reports the first characterisation of an [FeFe]-hydrogenase from a Clostridium perfringens strain previously isolated in our laboratory from a pilot-scale bio-hydrogen plant, that efficiently produces H2 from waste biomasses. On the basis of sequence analysis, the enzyme is a monomer formed by four domains hosting various iron-sulphur centres involved in electron transfer and the catalytic centre H-cluster. After recombinant expression in E. coli, the purified protein catalyses H2 evolution at high rate of 1645±16 s−1. The optimal conditions for catalysis are in the pH range 6.5-8.0 and at the temperature of 50°C. EPR spectroscopy showed that the H-cluster of the oxidised enzyme displays a spectrum coherent with the Hox state, while the CO inhibited enzyme has a spectrum coherent with the Hox-CO state. FTIR spectroscopy showed that the purified enzyme is composed of a mixture of redox states, with a prevalence of the Hox; upon reduction with H2, vibrational modes assigned to the Hred state were more abundant, while binding of exogenous CO resulted in a spectrum assigned to the Hox-CO state. The spectroscopic features observed are similar to those of the [FeFe]-hydrogenases class, but relevant differences were observed given the different protein environment hosting the H-cluster.

Isolation and characterisation of a new [FeFe]-hydrogenase from Clostridium perfringens.

MORRA, SIMONE;MAURELLI, SARA;GILARDI, Gianfranco;VALETTI, Francesca
Last
2016-01-01

Abstract

This paper reports the first characterisation of an [FeFe]-hydrogenase from a Clostridium perfringens strain previously isolated in our laboratory from a pilot-scale bio-hydrogen plant, that efficiently produces H2 from waste biomasses. On the basis of sequence analysis, the enzyme is a monomer formed by four domains hosting various iron-sulphur centres involved in electron transfer and the catalytic centre H-cluster. After recombinant expression in E. coli, the purified protein catalyses H2 evolution at high rate of 1645±16 s−1. The optimal conditions for catalysis are in the pH range 6.5-8.0 and at the temperature of 50°C. EPR spectroscopy showed that the H-cluster of the oxidised enzyme displays a spectrum coherent with the Hox state, while the CO inhibited enzyme has a spectrum coherent with the Hox-CO state. FTIR spectroscopy showed that the purified enzyme is composed of a mixture of redox states, with a prevalence of the Hox; upon reduction with H2, vibrational modes assigned to the Hred state were more abundant, while binding of exogenous CO resulted in a spectrum assigned to the Hox-CO state. The spectroscopic features observed are similar to those of the [FeFe]-hydrogenases class, but relevant differences were observed given the different protein environment hosting the H-cluster.
2016
63
3
305
311
http://onlinelibrary.wiley.com/doi/10.1002/bab.1382/abstract
Bio-hydrogen; Clostridium perfringens; [FeFe]-hydrogenases; recombinant expression.; iron-sulphur centres; H-cluster
Morra S; Mongili Beatrice; Maurelli S; Gilardi G; Valetti F.
File in questo prodotto:
File Dimensione Formato  
post print BAB Morra 2015.pdf

Open Access dal 01/06/2017

Tipo di file: POSTPRINT (VERSIONE FINALE DELL’AUTORE)
Dimensione 243.88 kB
Formato Adobe PDF
243.88 kB Adobe PDF Visualizza/Apri

I documenti in IRIS sono protetti da copyright e tutti i diritti sono riservati, salvo diversa indicazione.

Utilizza questo identificativo per citare o creare un link a questo documento: https://hdl.handle.net/2318/1509236
Citazioni
  • ???jsp.display-item.citation.pmc??? 4
  • Scopus 9
  • ???jsp.display-item.citation.isi??? 9
social impact