Esterases receive special attention because of their wide distribution in biological systems and environments and their importance for physiology and chemical synthesis. The prediction of esterases' substrate promiscuity level from sequence data and the molecular reasons why certain such enzymes are more promiscuous than others remain to be elucidated. This limits the surveillance of the sequence space for esterases potentially leading to new versatile biocatalysts and new insights into their role in cellular function. Here, we performed an extensive analysis of the substrate spectra of 145 phylogenetically and environmentally diverse microbial esterases, when tested with 96 diverse esters. We determined the primary factors shaping their substrate range by analyzing substrate range patterns in combination with structural analysis and protein-ligand simulations. We found a structural parameter that helps rank (classify) the promiscuity level of esterases from sequence data at 94% accuracy. This parameter, the active site effective volume, exemplifies the topology of the catalytic environment by measuring the active site cavity volume corrected by the relative solvent accessible surface area (SASA) of the catalytic triad. Sequences encoding esterases with active site effective volumes (cavity volume/SASA) above a threshold show greater substrate spectra, which can be further extended in combination with phylogenetic data. This measure provides also a valuable tool for interrogating substrates capable of being converted. This measure, found to be transferred to phosphatases of the haloalkanoic acid dehalogenase superfamily and possibly other enzymatic systems, represents a powerful tool for low-cost bioprospecting for esterases with broad substrate ranges, in large scale sequence data sets.
SummaryRecent reports have suggested that the establishment of industrially relevant enzyme collections from environmental genomes has become a routine procedure. Across the studies assessed, a mean number of approximately 44 active clones were obtained in an average size of approximately 53 000 clones tested using naïve screening protocols. This number could be significantly increased in shorter times when novel metagenome enzyme sequences obtained by direct sequencing are selected and subjected to high‐throughput expression for subsequent production and characterization. The pre‐screening of clone libraries by naïve screens followed by the pyrosequencing of the inserts allowed for a 106‐fold increase in the success rate of identifying genes encoding enzymes of interest. However, a much longer time, usually on the order of years, is needed from the time of enzyme identification to the establishment of an industrial process. If the hit frequency for the identification of enzymes performing at high turnover rates under real application conditions could be increased while still covering a high natural diversity, the very expensive and time‐consuming enzyme optimization phase would likely be significantly shortened. At this point, it is important to review the current knowledge about the success of fine‐tuned naïve‐ and sequence‐based screening protocols for enzyme selection and to describe the environments worldwide that have already been subjected to enzyme screen programmes through metagenomic tools. Here, we provide such estimations and suggest the current challenges and future actions needed before environmental enzymes can be successfully introduced into the market.
While changes in gut microbial populations have been described in human immuno-deficiency virus (HIV)-infected patients undergoing antiretroviral therapy (ART), the mechanisms underlying the contributions of gut bacteria and their molecular agents (metabolites and proteins) to immune recovery remain unexplored. To study this, we examined the active fraction of the gut microbiome, through examining protein synthesis and accumulation of metabolites inside gut bacteria and in the bloodstream, in 8 healthy controls and 29 HIV-infected individuals (6 being longitudinally studied). We found that HIV infection is associated to dramatic changes in the active set of gut bacteria simultaneously altering the metabolic outcomes. Effects were accentuated among immunological ART responders, regardless diet, subject characteristics, clinical variables other than immune recovery, the duration and type of ART and sexual preferences. The effect was found at quantitative levels of several molecular agents and active bacteria which were herein identified and whose abundance correlated with HIV immune pathogenesis markers. Although, we cannot rule out the possibility that some changes are partially a random consequence of the disease status, our data suggest that most likely reduced inflammation and immune recovery is a joint solution orchestrated by both the active fraction of the gut microbiota and the host.
Multiple factors have been shown to alter intestinal microbial diversity. It remains to be seen, however, how multiple collective pressures impact the activity in the gut environment and which, if any, is positioned as a dominant driving factor determining the final metabolic outcomes. Here, we describe the results of a metabolome-wide scan of gut microbiota in 18 subjects with systemic lupus erythematosus (SLE) and 17 healthy control subjects and demonstrate a statistically significant difference (p < 0.05) between the two groups. Healthy controls could be categorized (p < 0.05) based on their body mass index (BMI), whereas individuals with SLE could not. We discuss the prevalence of SLE compared with BMI as the dominant factor that regulates gastrointestinal microbial metabolism and provide plausible explanatory causes. Our results uncover novel perspectives with clinical relevance for human biology. In particular, we rank the importance of various pathophysiologies for gut homeostasis.
The microbiomes in the gastrointestinal tract (GIT) of individuals receiving antibiotics and those in obese subjects undergo compositional shifts, the metabolic effects and linkages of which are not clearly understood. Herein, we set to gain insight into these effects, particularly with regard to carbohydrate metabolism, and to contribute to unravel the underlying mechanisms and consequences for health conditions. We measured the activity level of GIT carbohydrate-active enzymes toward 23 distinct sugars in adults patients (n = 2) receiving 14-d β-lactam therapy and in obese (n = 7) and lean (n = 5) adolescents. We observed that both 14 d antibiotic-treated and obese subjects showed higher and less balanced sugar anabolic capacities, with 40% carbohydrates being preferentially processed as compared with non-treated and lean patients. Metaproteome-wide metabolic reconstructions confirmed that the impaired utilization of sugars propagated throughout the pentose phosphate metabolism, which had adverse consequences for the metabolic status of the GIT microbiota. The results point to an age-independent positive association between GIT glycosidase activity and the body mass index, fasting blood glucose and insulin resistance (r2 ≥ 0.95). Moreover, antibiotics altered the active fraction of enzymes controlling the thickness, composition and consistency of the mucin glycans. Our data and analyses provide biochemical insights into the effects of antibiotic usage on the dynamics of the GIT microbiota and pin-point presumptive links to obesity. The knowledge and the hypotheses generated herein lay a foundation for subsequent, systematic research that will be paramount for the design of “smart” dietary and therapeutic interventions to modulate host-microbe metabolic co-regulation in intestinal homeostasis.
Two of the largest crude oil-polluted areas in the world are the semi-enclosed Mediterranean and Red Seas, but the effect of chronic pollution remains incompletely understood on a large scale. We compared the influence of environmental and geographical constraints and anthropogenic forces (hydrocarbon input) on bacterial communities in eight geographically separated oil-polluted sites along the coastlines of the Mediterranean and Red Seas. The differences in community compositions and their biodegradation potential were primarily associated (P < 0.05) with both temperature and chemical diversity. Furthermore, we observed a link between temperature and chemical and biological diversity that was stronger in chronically polluted sites than in pristine ones where accidental oil spills occurred. We propose that low temperature increases bacterial richness while decreasing catabolic diversity and that chronic pollution promotes catabolic diversification. Our results further suggest that the bacterial populations in chronically polluted sites may respond more promptly in degrading petroleum after accidental oil spills.
The esterases and lipases from the ␣/ hydrolase superfamily exhibit an enormous sequence diversity, fold plasticity, and activities. Here, we present the comprehensive sequence and biochemical analyses of seven distinct esterases and lipases from the metagenome of Lake Arreo, an evaporite karstic lake in Spain (42°46=N, 2°59=W; altitude, 655 m). Together with oligonucleotide usage patterns and BLASTP analysis, our study of esterases/lipases mined from Lake Arreo suggests that its sediment contains moderately halophilic and cold-adapted proteobacteria containing DNA fragments of distantly related plasmids or chromosomal genomic islands of plasmid and phage origins. This metagenome encodes esterases/lipases with broad substrate profiles (tested over a set of 101 structurally diverse esters) and habitat-specific characteristics, as they exhibit maximal activity at alkaline pH (8.0 to 8.5) and temperature of 16 to 40°C, and they are stimulated (1.5 to 2.2 times) by chloride ions (0.1 to 1.2 M), reflecting an adaptation to environmental conditions. Our work provides further insights into the potential significance of the Lake Arreo esterases/lipases for biotechnology processes (i.e., production of enantiomers and sugar esters), because these enzymes are salt tolerant and are active at low temperatures and against a broad range of substrates. As an example, the ability of a single protein to hydrolyze triacylglycerols, (non)halogenated alkyl and aryl esters, cinnamoyl and carbohydrate esters, lactones, and chiral epoxides to a similar extent was demonstrated. E sterases and lipases from the ␣/ hydrolase family have received considerable attention, because they are widely distributed within the microbial communities operating in most of environments where they have important physiological functions (1) and because they are one of the most important groups of biocatalysts for biotechnological applications (2-4). Upon searching the list of genes using Pfam (the protein family database [5]) from the approximately 140 metagenomic projects in various stages of sequencing on the GOLD website (Genomes OnLine Database; http://www.genomesonline.org/) and the available sequences of esterases and lipases, more than 72,000 predicted esterases/lipases of the ␣/ hydrolase superfamily were retrieved, which revealed the richness of uncultured biodiversity (6), to provide wide collections of such biocatalysts. This is one of the largest protein families with available sequences. In relation to the cultivation-independent methods used to identify them, it should be highlighted that sequence-based metagenomics only provide the presumptive compositional and functional blueprint represented in the community genome (7,8), but at the same time, this method causes serious problems regarding both sequencing errors (9) and the erroneous assignment of substrate specificity (10). In contrast, the activity-directed techniques have been shown to provide a direct view of known or new protein families and functionalities (for examples, see referen...
scite is a Brooklyn-based organization that helps researchers better discover and understand research articles through Smart Citations–citations that display the context of the citation and describe whether the article provides supporting or contrasting evidence. scite is used by students and researchers from around the world and is funded in part by the National Science Foundation and the National Institute on Drug Abuse of the National Institutes of Health.
hi@scite.ai
10624 S. Eastern Ave., Ste. A-614
Henderson, NV 89052, USA
Copyright © 2024 scite LLC. All rights reserved.
Made with 💙 for researchers
Part of the Research Solutions Family.