Exploring the Molecular Basis for Substrate Affinity and Structural Stability in Bacterial GH39 β-Xylosidases.

The glycoside hydrolase family 39 (GH39) is a functionally expanding family with limited understanding about the molecular basis for substrate specificity and extremophilicity. In this work, we demonstrate the key role of the positive-subsite region in modulating substrate affinity and how the lack of a C-terminal extension impacts on oligomerization and structural stability of some GH39 members. The crystallographic and SAXS structures of a new GH39 member from the phytopathogen support the importance of an extended C-terminal to promote oligomerization as a molecular strategy to enhance thermal stability.

A single P115Q mutation modulates specificity in the Corynebacterium pseudotuberculosis arginine repressor.

The arginine repressor (ArgR) regulates the expression of genes involved in arginine biosynthesis. Upon attaining a threshold concentration of arginine in the cytoplasm, the trimeric C-terminal domain of ArgR binds three arginines in a shallow surface cleft and subsequently hexamerizes forming a dimer of trimers containing six Arg co-repressor molecules which are buried at the subunit interfaces. The N-terminal domains of this complex bind to the DNA promoter thereby interrupting the transcription of the genes related to Arg biosynthesis.

Spatially remote motifs cooperatively affect substrate preference of a ruminal GH26-type endo-β-1,4-mannanase.

β-Mannanases from the glycoside hydrolase 26 (GH26) family are retaining hydrolases that are active on complex heteromannans and whose genes are abundant in rumen metagenomes and metatranscriptomes. These enzymes can exhibit distinct modes of substrate recognition and are often fused to carbohydrate-binding modules (CBMs), resulting in a molecular puzzle of mechanisms governing substrate preference and mode of action that has not yet been pieced together. In this study, we recovered a novel GH26 enzyme with a CBM35 module linked to its N terminus (CrMan26) from a cattle rumen metatranscriptome.

Structure-guided design combined with evolutionary diversity led to the discovery of the xylose-releasing exo-xylanase activity in the glycoside hydrolase family 43.

Rational design is an important tool for sculpting functional and stability properties of proteins and its potential can be much magnified when combined with in vitro and natural evolutionary diversity. Herein, we report the structure-guided design of a xylose-releasing exo-β-1,4-xylanase from an inactive member of glycoside hydrolase family 43 (GH43). Structural analysis revealed a nonconserved substitution (Lys ) that results in the disruption of the hydrogen bond network that supports catalysis.

The mechanism by which a distinguishing arabinofuranosidase can cope with internal di-substitutions in arabinoxylans.

Background: Arabinoxylan is an abundant polysaccharide in industrially relevant biomasses such as sugarcane, corn stover and grasses. However, the arabinofuranosyl di-substitutions that decorate the xylan backbone are recalcitrant to most known arabinofuranosidases (Abfs).

Results: In this work, we identified a novel GH51 Abf (Abf51) that forms trimers in solution and can cope efficiently with both mono- and di-substitutions at terminal or internal xylopyranosyl units of arabinoxylan.

Structural basis of exo-β-mannanase activity in the GH2 family.

The classical microbial strategy for depolymerization of β-mannan polysaccharides involves the synergistic action of at least two enzymes, endo-1,4-β-mannanases and β-mannosidases. In this work, we describe the first exo-β-mannanase from the GH2 family, isolated from pv. (XacMan2A), which can efficiently hydrolyze both manno-oligosaccharides and β-mannan into mannose.

A novel β-glucosidase isolated from the microbial metagenome of Lake Poraquê (Amazon, Brazil).

The Amazon region holds most of the biological richness of Brazil. Despite their ecological and biotechnological importance, studies related to microorganisms from this region are limited. Metagenomics leads to exciting discoveries, mainly regarding non-cultivable microorganisms.

Cloning, expression, purification, crystallization and preliminary X-ray diffraction analysis of the mitochondrial tryparedoxin peroxidase from Leishmania braziliensis.

Tryparedoxin peroxidase (TXNPx) is an essential constituent of the main enzymatic scavenger system for reactive oxygen species (ROS) in trypanosomatids. Genetic studies have demonstrated the importance of this system for the development and virulence of these parasites, representing a potential target for the discovery of new trypanocidal drugs. In this work, the mitochondrial TXNPx from Leishmania braziliensis was cloned, overexpressed, purified and crystallized.