RESUMO
Extreme acidophilic bacteria like Leptospirillum sp. require an efficient enzyme system to counteract strong oxygen stress conditions in their natural habitat. The genome of Leptospirillum sp. CF-1 encodes the thioredoxin-fold protein TFP2, which exhibits a high structural similarity to the thioredoxin domain of E. coli CnoX. CnoX from Escherichia coli is a chaperedoxin that protects protein substrates from oxidative stress conditions using its holdase function and a subsequent transfer to foldase chaperones for refolding. Recombinantly produced and purified Leptospirillum sp. TFP2 possesses both thioredoxin and chaperone holdase activities in vitro. It can be reduced by thioredoxin reductase (TrxR). The tfp2 gene co-locates with genes for the chaperone foldase GroES/EL on the chromosome. The "tfp2 cluster" (ctpA-groES-groEL-hyp-tfp2-recN) was found between 1.9 and 8.8-fold transcriptionally up-regulated in response to 1 mM hydrogen peroxide (H2O2). Leptospirillum sp. tfp2 heterologously expressed in E. coli wild type and cnoX mutant strains lead to an increased tolerance of these E. coli strains to H2O2 and significantly reduced intracellular protein aggregates. Finally, a proteomic analysis of protein aggregates produced in E. coli upon exposition to oxidative stress with 4 mM H2O2, showed that Leptospirillum sp. tfp2 expression caused a significant decrease in the aggregation of 124 proteins belonging to fifteen different metabolic categories. These included several known substrates of DnaK and GroEL/ES. These findings demonstrate that Leptospirillum sp. TFP2 is a chaperedoxin-like protein, acting as a key player in the control of cellular proteostasis under highly oxidative conditions that prevail in extreme acidic environments.
Assuntos
Proteínas de Bactérias , Estresse Oxidativo , Tiorredoxinas , Proteínas de Bactérias/metabolismo , Proteínas de Bactérias/genética , Tiorredoxinas/metabolismo , Tiorredoxinas/genética , Escherichia coli/metabolismo , Escherichia coli/genética , Chaperonas Moleculares/metabolismo , Chaperonas Moleculares/genética , Agregados Proteicos , Peróxido de Hidrogênio/farmacologia , Peróxido de Hidrogênio/metabolismo , Regulação Bacteriana da Expressão GênicaRESUMO
We describe the complete genome sequence of Leptospirillum sp. group II strain CF-1, an acidophilic bioleaching bacterium isolated from an acid mine drainage (AMD). This work provides data to gain insights about adaptive response of Leptospirillum spp. to the extreme conditions of bioleaching environments.
Assuntos
Bactérias/genética , Genoma Bacteriano/genética , Microbiologia Ambiental , Poluentes Químicos da ÁguaRESUMO
Glutamyl-tRNA reductase (GluTR) is the first enzyme committed to tetrapyrrole biosynthesis by the C(5)-pathway. This enzyme transforms glutamyl-tRNA into glutamate-1-semi-aldehyde, which is then transformed into 5-amino levulinic acid by the glutamate-1-semi-aldehyde 2,1-aminomutase. Binding of heme to GluTR seems to be relevant to regulate the enzyme function. Recombinant GluTR from Acidithiobacillus ferrooxidans an acidophilic bacterium that participates in bioleaching of minerals was expressed in Escherichia coli and purified as a soluble protein containing type b heme. Upon control of the cellular content of heme in E. coli, GluTR with different levels of bound heme was obtained. An inverse correlation between the activity of the enzyme and the level of bound heme to GluTR suggested a control of the enzyme activity by heme. Heme bound preferentially to dimeric GluTR. An intact dimerization domain was essential for the enzyme to be fully active. We propose that the cellular levels of heme might regulate the activity of GluTR and ultimately its own biosynthesis.