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In mitochondrial oxidative phosphorylation, electron transfer from NADH or succinate to oxygen by a series of large protein complexes in the inner mitochondrial membrane (complexes I–IV) is coupled to the generation of an electrochemical proton gradient, the energy of which is utilized by complex V to generate ATP. In Euglena gracilis, a non-parasitic secondary green alga related to trypanosomes, these respiratory complexes totalize more than 40 Euglenozoa-specific subunits along with about 50 classical subunits described in other eukaryotes. In the present study the Euglena proton-pumping complexes I, III, and IV were purified from isolated mitochondria by a two-steps liquid chromatography approach. Their atypical subunit composition was further resolved and confirmed using a three-steps PAGE analysis coupled to mass spectrometry identification of peptides. The purified complexes were also observed by electron microscopy followed by single-particle analysis. Even if the overall structures of the three oxidases are similar to the structure of canonical enzymes (e.g. from mammals), additional atypical domains were observed in complexes I and IV: an extra domain located at the tip of the peripheral arm of complex I and a “helmet-like” domain on the top of the cytochrome c binding region in complex IV.

From the extracellular fluid of a novel strain of Bjerkandera sp., it was isolated, purified and identified the main enzyme responsible for Remazol Brilliant Blue R dye decolourisation. Such an enzyme is able to oxidise manganese, as well as veratryl alcohol and 2,6-dimethoxyphenol in manganese-independent reactions; hence, it can be included in the new group of versatile peroxidases. The molecular mass of said enzyme is ca. 45 kDa, and the N-terminal amino acid sequence obtained by Edman degradation is VAXPDGVNTA. The enzyme substrate range for oxidation of several phenolic and non-phenolic aromatic compounds was determined and the corresponding Michaelis–Menten kinetic constants calculated. Furthermore, spectrophotometric assays showing the Soret band and allowing observation of band shifts of the enzyme led to the conclusion that Bjerkandera strains may also synthesise at least two different versatile peroxidases, as happens with Pleurotus eryngii .

Bacterial resistance to beta-lactam antibiotics, a clinically worrying and recurrent problem, is often due to the production of beta-lactamases, enzymes that efficiently hydrolyze the amide bond of the beta-lactam nucleus. Imipenem and other carbapenems escape the activity of most active site serine beta-lactamases and have therefore become very popular drugs for antibacterial chemotherapy in the hospital environment. Their usefulness is, however, threatened by the appearance of new beta-lactamases that efficiently hydrolyze them. This study is focused on the structure and properties of two recently described class A carbapenemases, produced by Serratia marcescens and Enterobacter cloacae strains and leads to a better understanding of the specificity of beta-lactamases. In turn, this will contribute to the design of better antibacterial drugs. Three-dimensional models of the two class A carbapenemases were constructed by homology modeling. They suggested the presence, near the active site of the enzymes, of a disulfide bridge (C69-C238) whose existence was experimentally confirmed. Kinetic parameters were measured with the purified Sme-1 carbapenemase, and an attempt was made to explain its specific substrate profile by analyzing the structures of minimized Henri-Michaelis complexes and comparing them to those obtained for the "classical" TEM-1 beta-lactamase. The peculiar substrate profile of the carbapenemases appears to be strongly correlated with the presence of the disulfide bridge between C69 and C238.