Recent advances in the pretreatment of microalgal and lignocellulosic biomass: A comprehensive review.

Microalgal and lignocellulosic biomass is the most sumptuous renewable bioresource raw material existing on earth. Recently, the bioconversion of biomass into biofuels have received significant attention replacing fossil fuels. Pretreatment of biomass is a critical process in the conversion due to the nature and structure of the biomass cell wall that is complex. Although green technologies for biofuel production are advancing, the productivity and yield from these techniques are low. Over the past years, various pretreatment techniques have been developed and successfully employed to improve the technology. This paper presents an in-depth review of the recent advancement of pretreatment methods focusing on microalgal and lignocellulosic biomass. The technological approaches involving physical, chemical, biological and other latest pretreatment methods are reviewed.

Revealing solvent-dependent folding behavior of mycolic acids from Mycobacterium tuberculosis by advanced simulation analysis.

Mycobacterium tuberculosis remains a persistent pathogen, partly due to its lipid rich cell wall, of which mycolic acids (MAs) are a major component. The fluidity and conformational flexibilities of different MAs in the bacterial cell wall significantly influence its properties, function, and observed pathogenicity; thus, a proper conformational description of different MAs in different environments (e.g., in vacuum, in solution, in monolayers) can inform about their potential role in the complex setup of the bacterial cell wall. Previously, we have shown that molecular dynamics (MD) simulations of MA folding in vacuo can be used to characterize MA conformers in seven groupings relating to bending at the functional groups (W, U and Z-conformations). Providing a new OPLS-based forcefield parameterization for the critical cyclopropyl group of MAs and extensive simulations in explicit solvents (TIP4P water, hexane), we now present a more complete picture of MA folding properties together with improved simulation analysis techniques. We show that the 'WUZ' distance-based analysis can be used to pinpoint conformers with hairpin bends at the functional groups, with these conformers constituting only a fraction of accessible conformations. Applying principle component analysis (PCA) and refinement using free energy landscapes (FELs), we are able to discriminate a complete and unique set of conformational preferences for representative alpha-, methoxy- and keto-MAs, with overall preference for folded conformations. A control backbone-MA without any mero-chain functional groups showed significantly less folding in the mero-chain, confirming the role of functionalization in directing folding. Keto-MA showed the highest percentage of WUZ-type conformations and, in particular, a tendency to fold at its alpha-methyl trans-cyclopropane group, in agreement with results from Villeneuve et al. MAs demonstrate similar folding in vacuum and water, with a majority of folded conformations around the W-conformation, although the molecules are more flexible in vacuum than in water. Exchange between conformations, with a disperse distribution that includes unfolded conformers, is common in hexane for all MAs, although with more organization for Keto-MA. Globular, folded conformations are newly defined and may be specifically relevant in biofilms. Graphical abstract Through advanced simulation analysis, including principle component analysis and free energy landscapes, we reveal detailed physical insights into the solvent-dependant folding behavior of mycolic acids from M. tb.

Rational Design of Thermostable Carbonic Anhydrase Mutants Using Molecular Dynamics Simulations.

The stability of enzymes is critical for their application in industrial processes, which generally require different conditions from the natural enzyme environment. Both rational and random protein engineering approaches have been used to increase stability, with the latter requiring extensive experimental effort for the screening of variants. Moreover, some general rules addressing the molecular origin of protein thermostability have been established. Herein, we demonstrate the use of molecular dynamics simulations to gain molecular level understanding of protein thermostability and to engineer stabilizing mutations. Carbonic anhydrase (CA) is an enzyme with a high potential for biotechnological carbon capture applications, provided it can be engineered to withstand the high temperature process environments, inevitable in most gas treatment units. In this study, we used molecular dynamics simulations at 343, 353, and 363 K to study the relationship between structure flexibility and thermostability in bacterial α-CAs and applied this knowledge to the design of mutants with increased stability. The most thermostable α-CA known, TaCA from Thermovibrio ammonificans, had the most rigid structure during molecular dynamics simulations, but also showed regions with high flexibility. The most flexible amino acids in these regions were identified from root mean square fluctuation (RMSF) studies, and stabilizing point mutations were predicted based on their capacity to improve the calculated free energy of unfolding. Disulfide bonds were also designed at sites with suitable geometries and selected based on their location at flexible sites, assessed by B-factor calculation. Molecular dynamics simulations allowed the identification of five mutants with lower RMSF of the overall structure at 400 K, compared to wild-type TaCA. Comparison of free-energy landscapes between wild-type TaCA and the most promising mutants, Pro165Cys-Gln170Cys and Asn140Gly, showed an increased conformational stability of the mutants at 400 K.