Mechanism Of Naphthalene Degradation And Chromate Reduction In Pseudomonas Brassicacearum LZ-4

Hexavalent chromium and naphthalene have a huge threat to human health.In this paper,four proteins(NahAa,NahG,NahE and NahR),which were involved in naphthalene degradation in Pseudomonas aeruginosa strain LZ-4 were studied.Previous study showed that strain LZ-4 could degrade naphthalene and reduce hexavalent chromium simultaneously,while no genes related to Cr(?)reduction were found in its whole genome sequencing data.So these four proteins,which crystallization structures were unclear,were studied.In these four proteins,NahAa(naphthalene 1,2-dioxygenase ferredoxin reductase)take part in the first step of naphthalene degradation,NahE(trans-o-hydroxybenzylidenepyruvate hydratase-aldolase)is responsible for salicylaldehyde formation,NahG catalyze salicylic acid reduced to catechol,and NahR is the transcriptional regulation factor of naphthalene degradation.So,NahAa and NahG were chosen to determine the ability of Cr(?)reduction,and crystal structure analysis of NahE and the catalytic function of NahR protein were performed after expression and purification of these proteins through Ni2+-nitrilotriacetate affinity method.Resluts showed that 10 ?M NahAa was able to reduce 69.21% hexavalent chromium in 1 min,and 10 ?M NahG could react with 79.17% hexavalent chromium in the system when the sodium salicylate was added as the common substrate in 1 min.After purification of NahE,the crystallization of the protein was carried out and a total of 69 primary screening conditions were confirmed.Then optimizing crystallographic conditions were operated and the results of optimization were not very well.Therefore,the primary crystals were used for the follow-up experiments.The purified method of denaturation and renaturation was used for purification of NahR after several attempts.Then,a small amount of NahR protein was purified.This study demonstrated that NahAa and NahG not only take part in naphthalene degradation,but also were involved in Cr(?)reduction in strain LZ-4,and also provide useful information for biodegradation of naphthalene and Cr(?)by using microbes or enzyme.