Role of Sinorhizobium meliloti and Escherichia coli Long-Chain Acyl-CoA Synthetase FadD in Long-Term Survival.

FadD is an acyl-coenzyme A (CoA) synthetase specific for long-chain fatty acids (LCFA). Strains mutated in fadD cannot produce acyl-CoA and thus cannot grow on exogenous LCFA as the sole carbon source. Mutants in the fadD (smc02162) of Sinorhizobium meliloti are unable to grow on oleate as the sole carbon source and present an increased surface motility and accumulation of free fatty acids at the entry of the stationary phase of growth. In this study, we found that constitutive expression of the closest FadD homologues of S. meliloti, encoded by sma0150 and smb20650, could not revert any of the mutant phenotypes. In contrast, the expression of Escherichia coli fadD could restore the same functions as S. meliloti fadD. Previously, we demonstrated that FadD is required for the degradation of endogenous fatty acids released from membrane lipids. Here, we show that absence of a functional fadD provokes a significant loss of viability in cultures of E. coli and of S. meliloti in the stationary phase, demonstrating a crucial role of fatty acid degradation in survival capacity.

Functional Expression and One-Step Protein Purification of Manganese Peroxidase 1 (rMnP1) from Phanerochaete chrysosporium Using the E. coli-Expression System.

Manganese peroxidases (MnP) from the white-rot fungi Phanerochaete chrysosporium catalyse the oxidation of Mn2+ to Mn3+, a strong oxidizer able to oxidize a wide variety of organic compounds. Different approaches have been used to unravel the enzymatic properties and potential applications of MnP. However, these efforts have been hampered by the limited production of native MnP by fungi. Heterologous expression of MnP has been achieved in both eukaryotic and prokaryotic expression systems, although with limited production and many disadvantages in the process. Here we described a novel molecular approach for the expression and purification of manganese peroxidase isoform 1 (MnP1) from P. chrysosporium using an E. coli-expression system. The proposed strategy involved the codon optimization and chemical synthesis of the MnP1 gene for optimised expression in the E. coli T7 shuffle host. Recombinant MnP1 (rMnP1) was expressed as a fusion protein, which was recovered from solubilised inclusion bodies. rMnP1 was purified from the fusion protein using intein-based protein purification techniques and a one-step affinity chromatography. The designated strategy allowed production of an active enzyme able to oxidize guaiacol or Mn2+.

A Brief View of the Surface Membrane Proteins from Trypanosoma cruzi.

Trypanosoma cruzi is the causal agent of Chagas' disease which affects millions of people around the world mostly in Central and South America. T. cruzi expresses a wide variety of proteins on its surface membrane which has an important role in the biology of these parasites. Surface molecules of the parasites are the result of the environment to which the parasites are exposed during their life cycle. Hence, T. cruzi displays several modifications when they move from one host to another. Due to the complexity of this parasite's cell surface, this review presents some membrane proteins organized as large families, as they are the most abundant and/or relevant throughout the T. cruzi membrane.