MULTIPLE RESPIRATORY BURSTS AS A RESPONSE TO VERATRATE STRESS IN RHODOCOCCUS ERYTHROPOLIS CELLS

Abstract

Although Rhodococcus spp. strains are able to degrade methoxyphenols by enzymatic means, the contact with veratric acid (3,4-dimethoxybenzoic acid, hereafter called veratrate) is very stressful for the cells ofRhodococcus erythropolis DSM 1069 (Rh). Within 5min of contact veratrate in phosphate buffer, the emergence of many vacuoles was observed in the cell body and respiratory bursts, with violent endogenous oxygen uptake, took place several times during the 24h incubation. During these peaks (where the cells were in their MAX states), increased activity of NADH oxidase was noted, accompanied by maximal accumulation of vanillic and isovanillic acids (3-methoxy-4-hydroxybenzoic acid and 4-hydroxy-3-methoxybenzoic acid respectively, hereafter called vanillates) in the incubation medium, which appeared to be products of veratrate demethylation. At the troughs (cell in their MIN state), the vacuoles disappeared from the cell body, oxygen uptake was normal, and the pool of vanillates decreased while the veratrate level in the medium increased. The cells from MAX and MIN states reacted in opposite ways in the presence of either formaldehyde and GSH, or paraquate and cAMP. The NADH oxidase activity, measured as oxygen uptake against NADH in the membrane pellets of MAX and MIN stage cells, differed in their response to the exogenous presence of FAD, ATP, cAMP, catalase, GSH, H2O2and methoxyphenolic substrates. The periodic character of these events is described here. Co-operation between two multiprotein membrane complexes (NAD(P)H oxidase and 3-O/4-O-demethylases) in Rhodococcus erythropolis cells and their competition for two common substrates—NAD(P)H and O2—is proposed as an explanation for rhythmical nature of these reactions.