Analysis of phenanthrene catabolic pathway in Sphingomonas paucimobilis 20006FA using a proteomic approach

COPOTELLI, B; MARTINEZ, M; RODRIGUEZ, ME; MORELLI, I

Seattle, USA

Congreso; 13th International Symposium on Microbial Ecology (ISME-13).; 2010

International Microbial Ecology Society

In bioremediation applications, cellular physiology is an important issue which provides essential biological knowledge for the design and optimization of biodegradation processes. In order to study the phenanthrene catabolic pathway, the Sphingomonas paucimobilis 20006FA proteome was analyzed under two different growth substrate conditions, using glucose or phenanthrene as sole carbon source. Kinetics of growth, phenanthrene degradation, and 1-hydroxy-2-naphthoic acid production, a key metabolite of phenanthrene degradation, were monitored. Cells were harvested at the beginning of the stationary phase (96 h). The protein analysis was performed by two dimensional gel electrophoresis and mass spectrometry (MALDI-TOF/TOF MS). At the harvest time, a X % of phenanthrene was biodegraded, with the concomitant accumulation of .…ppm of 1-hydroxy-2-naphthoic acid. Soluble cellular protein fractions were analyzed using immobilized pH gradient (IPG) strips within the window of isoelectric point (pI) 4-7 and a molecular mass range of 6.5 to 200 kDa. Ten PAH-induced proteins were identified, including enzymes from the upper catabolic pathway of phenanthrene (extradiol dioxygenase, bencene dioxygenase, dihidrodiol dehydrogenase, 2-hydroxy-benzylpyruvate aldolase); and from the lower catabolic pathway of phenanthrene, all belonging to the meta cleavage pathway, (catecol 2,3 dioxigenase, 4-oxalocrotonate decarboxylase, 2- hydroxymuconic semialdehide hydrolasa, 2-hydroxypent-2,4-dienoate hydratase, 4-hydroxy-2-ovovalerte aldolase). The glutation S-transferase, a multifunctional enzyme involved in the cellular detoxification and excretion of a large variety of compounds, was also identified. This proteomic approach enabled the full description of the phenanthrene degradation pathway we have previously proposed by means of key intermediate metabolites identification.