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Thermophilic and Hyperthermophilic Microbes and Enzymes

A special issue of International Journal of Molecular Sciences (ISSN 1422-0067). This special issue belongs to the section "Molecular Microbiology".

Deadline for manuscript submissions: closed (31 August 2020) | Viewed by 22611

Special Issue Editors

Institute of Biochemistry and Cell Biology, National Research Council, Via Pietro Castellino 111, 80131 Naples, Italy
Interests: structure and function of thermostable enzymes; human enzymes and disease; proteomics; bioremediation and detoxification; biosensors; functional materials
Special Issues, Collections and Topics in MDPI journals
Biosciences Center, National Renewable Energy Laboratory, 15013 Denver West Parkway, Golden, CO 80401, USA
Interests: biomass deconstruction and conversion; enzyme engineering; cell free production of biochemicals
Special Issues, Collections and Topics in MDPI journals

Special Issue Information

Dear Colleagues,

Thermophilic and hyperthermophilic microbes represent a fascinating class of microorganisms, not only because of their resilience to thrive at elevated temperatures and in harsh environments, but also because of the enzymes that they harbor. These microorganisms fall mainly in the bacteria and archaea domains, and exist in many habitats including hot springs, hydrothermal vents, or volcanic ash sediments, among others. These habitats all exhibit thermophilic or hyperthermophilic temperatures, but can also be acidic, alkaline, or contain high levels of salts. In order to thrive in such harsh environments, these microorganisms have evolved robust enzymes that are able to function at peak activity in these harsh conditions. These microorganisms represent a rich source of enzymes with an increased stability, which are purposely modified by protein engineering and can excel in harsh industrial conditions, making them especially appealing for biotechnological applications. Sometimes, they display promiscous activities that represent a peculiar basis for evolution. Bioprospecting for enzymes in these microbes has gained popularity in the last two decades, as they also represent ideal templates and strategies for reengineering essential but less stable enzymes, catalyzing reactions that do not exist in thermophiles.

This Special Issue encourages original research articles, perspectives, and reviews on the topic of thermophilic and hyperthermophilic microbes and enzymes. Topics of interest include, but are not limited to, the following:

  • Bioprospecting
  • Enzyme characterization
  • Microbial phenotypes
  • Novel metabolic pathways
  • Novel metabolic enzymes
  • Bioremediation
  • Enzyme evolution and engineering
  • Proteomics approach
  • Microbial ecology
  • Microbial consortia
Welcome your submission to "Thermophilic and Hyperthermophilic Microbes and Enzymes 2.0".

Dr. Giuseppe Manco
Dr. Yannick J. Bomble
Guest Editors

Manuscript Submission Information

Manuscripts should be submitted online at www.mdpi.com by registering and logging in to this website. Once you are registered, click here to go to the submission form. Manuscripts can be submitted until the deadline. All submissions that pass pre-check are peer-reviewed. Accepted papers will be published continuously in the journal (as soon as accepted) and will be listed together on the special issue website. Research articles, review articles as well as short communications are invited. For planned papers, a title and short abstract (about 100 words) can be sent to the Editorial Office for announcement on this website.

Submitted manuscripts should not have been published previously, nor be under consideration for publication elsewhere (except conference proceedings papers). All manuscripts are thoroughly refereed through a single-blind peer-review process. A guide for authors and other relevant information for submission of manuscripts is available on the Instructions for Authors page. International Journal of Molecular Sciences is an international peer-reviewed open access semimonthly journal published by MDPI.

Please visit the Instructions for Authors page before submitting a manuscript. There is an Article Processing Charge (APC) for publication in this open access journal. For details about the APC please see here. Submitted papers should be well formatted and use good English. Authors may use MDPI's English editing service prior to publication or during author revisions.

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Published Papers (7 papers)

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Research

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10 pages, 2688 KiB  
Article
Altered Cofactor Preference of Thermostable StDAPDH by a Single Mutation at K159
by Xiuzhen Gao, Qinyuan Ma, Huihui Song, Xinming Sun, Zhiyun Li and Mingfei Liu
Int. J. Mol. Sci. 2020, 21(5), 1788; https://0-doi-org.brum.beds.ac.uk/10.3390/ijms21051788 - 05 Mar 2020
Cited by 2 | Viewed by 2647
Abstract
D-amino acid production from 2-keto acid by reductive amination is an attractive pathway because of its high yield and environmental safety. StDAPDH, a meso-diaminopimelate dehydrogenase (meso-DAPDH) from Symbiobacterium thermophilum, was the first meso-DAPDH to show amination of 2-keto [...] Read more.
D-amino acid production from 2-keto acid by reductive amination is an attractive pathway because of its high yield and environmental safety. StDAPDH, a meso-diaminopimelate dehydrogenase (meso-DAPDH) from Symbiobacterium thermophilum, was the first meso-DAPDH to show amination of 2-keto acids. Furthermore, StDAPDH shows excellent thermostability compared to other meso-DAPDHs. However, the cofactor of StDAPDH is NADP(H), which is less common than NAD(H) in industrial applications. Therefore, cofactor engineering for StDAPDH is needed. In this study, the highly conserved cofactor binding sites around the adenosine moiety of NADPH were targeted to determine cofactor specificity. Lysine residues within a loop were found to be critical for the cofactor specificity of StDAPDH. Replacement of lysine with arginine resulted in the activity of pyruvic acid with NADH as the cofactor. The affinity of K159R to pyruvic acid was equal with NADH or NADPH as the cofactor, regardless of the mutation. Molecular dynamics simulations revealed that the large steric hindrance of arginine and the interaction of the salt bridge between NADH and arginine may have restricted the free movement of NADH, which prompted the formation of a stable active conformation of mutant K159R. These results provide further understanding of the catalytic mechanism of StDAPDH and guidance for the cofactor engineering of StDAPDH. Full article
(This article belongs to the Special Issue Thermophilic and Hyperthermophilic Microbes and Enzymes)
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26 pages, 3797 KiB  
Article
Structural and Functional Characterization of New SsoPox Variant Points to the Dimer Interface as a Driver for the Increase in Promiscuous Paraoxonase Activity
by Yoko Suzumoto, Orly Dym, Giovanni N. Roviello, Franz Worek, Joel L. Sussman and Giuseppe Manco
Int. J. Mol. Sci. 2020, 21(5), 1683; https://0-doi-org.brum.beds.ac.uk/10.3390/ijms21051683 - 01 Mar 2020
Cited by 11 | Viewed by 2821
Abstract
Increasing attention is more and more directed toward the thermostable Phosphotriesterase-Like-Lactonase (PLL) family of enzymes, for the efficient and reliable decontamination of toxic nerve agents. In the present study, the DNA Staggered Extension Process (StEP) technique was utilized to obtain new variants of [...] Read more.
Increasing attention is more and more directed toward the thermostable Phosphotriesterase-Like-Lactonase (PLL) family of enzymes, for the efficient and reliable decontamination of toxic nerve agents. In the present study, the DNA Staggered Extension Process (StEP) technique was utilized to obtain new variants of PLL enzymes. Divergent homologous genes encoding PLL enzymes were utilized as templates for gene recombination and yielded a new variant of SsoPox from Saccharolobus solfataricus. The new mutant, V82L/C258L/I261F/W263A (4Mut) exhibited catalytic efficiency of 1.6 × 105 M−1 s−1 against paraoxon hydrolysis at 70°C, which is more than 3.5-fold and 42-fold improved in comparison with C258L/I261F/W263A (3Mut) and wild type SsoPox, respectively. 4Mut was also tested with chemical warfare nerve agents including tabun, sarin, soman, cyclosarin and VX. In particular, 4Mut showed about 10-fold enhancement in the hydrolysis of tabun and soman with respect to 3Mut. The crystal structure of 4Mut has been solved at the resolution of 2.8 Å. We propose that, reorganization of dimer conformation that led to increased central groove volume and dimer flexibility could be the major determinant for the improvement in hydrolytic activity in the 4Mut. Full article
(This article belongs to the Special Issue Thermophilic and Hyperthermophilic Microbes and Enzymes)
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18 pages, 1515 KiB  
Article
Metabolic Adaptation to Sulfur of Hyperthermophilic Palaeococcus pacificus DY20341T from Deep-Sea Hydrothermal Sediments
by Xiang Zeng, Xiaobo Zhang and Zongze Shao
Int. J. Mol. Sci. 2020, 21(1), 368; https://0-doi-org.brum.beds.ac.uk/10.3390/ijms21010368 - 06 Jan 2020
Cited by 6 | Viewed by 4388
Abstract
The hyperthermo-piezophilic archaeon Palaeococcus pacificus DY20341T, isolated from East Pacific hydrothermal sediments, can utilize elemental sulfur as a terminal acceptor to simulate growth. To gain insight into sulfur metabolism, we performed a genomic and transcriptional analysis of Pa. pacificus DY20341T [...] Read more.
The hyperthermo-piezophilic archaeon Palaeococcus pacificus DY20341T, isolated from East Pacific hydrothermal sediments, can utilize elemental sulfur as a terminal acceptor to simulate growth. To gain insight into sulfur metabolism, we performed a genomic and transcriptional analysis of Pa. pacificus DY20341T with/without elemental sulfur as an electron acceptor. In the 2001 protein-coding sequences of the genome, transcriptomic analysis showed that 108 genes increased (by up to 75.1 fold) and 336 genes decreased (by up to 13.9 fold) in the presence of elemental sulfur. Palaeococcus pacificus cultured with elemental sulfur promoted the following: the induction of membrane-bound hydrogenase (MBX), NADH:polysulfide oxidoreductase (NPSOR), NAD(P)H sulfur oxidoreductase (Nsr), sulfide dehydrogenase (SuDH), connected to the sulfur-reducing process, the upregulation of iron and nickel/cobalt transfer, iron–sulfur cluster-carrying proteins (NBP35), and some iron–sulfur cluster-containing proteins (SipA, SAM, CobQ, etc.). The accumulation of metal ions might further impact on regulators, e.g., SurR and TrmB. For growth in proteinous media without elemental sulfur, cells promoted flagelin, peptide/amino acids transporters, and maltose/sugar transporters to upregulate protein and starch/sugar utilization processes and riboflavin and thiamin biosynthesis. This indicates how strain DY20341T can adapt to different living conditions with/without elemental sulfur in the hydrothermal fields. Full article
(This article belongs to the Special Issue Thermophilic and Hyperthermophilic Microbes and Enzymes)
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12 pages, 2260 KiB  
Article
Occurrence of Thermophilic Microorganisms in Different Full Scale Biogas Plants
by Ivan Kushkevych, Jiří Cejnar, Monika Vítězová, Tomáš Vítěz, Dani Dordević and Yannick J. Bomble
Int. J. Mol. Sci. 2020, 21(1), 283; https://0-doi-org.brum.beds.ac.uk/10.3390/ijms21010283 - 31 Dec 2019
Cited by 10 | Viewed by 2967
Abstract
Background: In recent years, various substrates have been tested to increase the sustainable production of biomethane. The effect of these substrates on methanogenesis has been investigated mainly in small volume fermenters and were, for the most part, focused on studying the diversity of [...] Read more.
Background: In recent years, various substrates have been tested to increase the sustainable production of biomethane. The effect of these substrates on methanogenesis has been investigated mainly in small volume fermenters and were, for the most part, focused on studying the diversity of mesophilic microorganisms. However, studies of thermophilic communities in large scale operating mesophilic biogas plants do not yet exist. Methods: Microbiological, biochemical, biophysical methods, and statistical analysis were used to track thermophilic communities in mesophilic anaerobic digesters. Results: The diversity of the main thermophile genera in eight biogas plants located in the Czech Republic using different input substrates was investigated. In total, 19 thermophilic genera were detected after 16S rRNA gene sequencing. The highest percentage (40.8%) of thermophiles was found in the Modřice biogas plant where the input substrate was primary sludge and biological sludge (50/50, w/w %). The smallest percentage (1.87%) of thermophiles was found in the Čejč biogas plant with the input substrate being maize silage and liquid pig manure (80/20, w/w %). Conclusions: The composition of the anaerobic consortia in anaerobic digesters is an important factor for the biogas plant operator. The present study can help characterizing the impact of input feeds on the composition of microbial communities in these plants. Full article
(This article belongs to the Special Issue Thermophilic and Hyperthermophilic Microbes and Enzymes)
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Review

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13 pages, 1098 KiB  
Review
DING Proteins Extend to the Extremophilic World
by Elena Porzio, Maria Rosaria Faraone Mennella and Giuseppe Manco
Int. J. Mol. Sci. 2021, 22(4), 2035; https://0-doi-org.brum.beds.ac.uk/10.3390/ijms22042035 - 18 Feb 2021
Cited by 1 | Viewed by 1747
Abstract
The DING proteins are ubiquitous in the three domains of life, from mesophiles to thermo- and hyperthermophiles. They belong to a family of more than sixty members and have a characteristic N-terminus, DINGGG, which is considered a “signature” of these proteins. Structurally, they [...] Read more.
The DING proteins are ubiquitous in the three domains of life, from mesophiles to thermo- and hyperthermophiles. They belong to a family of more than sixty members and have a characteristic N-terminus, DINGGG, which is considered a “signature” of these proteins. Structurally, they share a highly conserved phosphate binding site, and a three dimensional organization resembling the “Venus Flytrap”, both reminding the ones of PstS proteins. They have unusually high sequence conservation, even between distantly related species. Nevertheless despite that the genomes of most of these species have been sequenced, the DING gene has not been reported for all the relative characterized DING proteins. Identity of known DING proteins has been confirmed immunologically and, in some cases, by N-terminal sequence analysis. Only a few of the DING proteins have been purified and biochemically characterized. DING proteins are heterogeneous for their wide range of biological activities and some show different activities not always correlated with each other. Most of them have been originally identified for different biological properties, or rather for binding to phosphate and also to other ligands. Their involvement in pathologies is described. This review is an update of the most recent findings on old and new DING proteins. Full article
(This article belongs to the Special Issue Thermophilic and Hyperthermophilic Microbes and Enzymes)
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34 pages, 1146 KiB  
Review
Effect of Cultivation Parameters on Fermentation and Hydrogen Production in the Phylum Thermotogae
by Mariamichela Lanzilli, Nunzia Esercizio, Marco Vastano, Zhaohui Xu, Genoveffa Nuzzo, Carmela Gallo, Emiliano Manzo, Angelo Fontana and Giuliana d’Ippolito
Int. J. Mol. Sci. 2021, 22(1), 341; https://0-doi-org.brum.beds.ac.uk/10.3390/ijms22010341 - 30 Dec 2020
Cited by 18 | Viewed by 3877
Abstract
The phylum Thermotogae is composed of a single class (Thermotogae), 4 orders (Thermotogales, Kosmotogales, Petrotogales, Mesoaciditogales), 5 families (Thermatogaceae, Fervidobacteriaceae, Kosmotogaceae, Petrotogaceae, Mesoaciditogaceae), and 13 genera. They have been isolated from extremely hot environments whose characteristics are [...] Read more.
The phylum Thermotogae is composed of a single class (Thermotogae), 4 orders (Thermotogales, Kosmotogales, Petrotogales, Mesoaciditogales), 5 families (Thermatogaceae, Fervidobacteriaceae, Kosmotogaceae, Petrotogaceae, Mesoaciditogaceae), and 13 genera. They have been isolated from extremely hot environments whose characteristics are reflected in the metabolic and phenotypic properties of the Thermotogae species. The metabolic versatility of Thermotogae members leads to a pool of high value-added products with application potentials in many industry fields. The low risk of contamination associated with their extreme culture conditions has made most species of the phylum attractive candidates in biotechnological processes. Almost all members of the phylum, especially those in the order Thermotogales, can produce bio-hydrogen from a variety of simple and complex sugars with yields close to the theoretical Thauer limit of 4 mol H2/mol consumed glucose. Acetate, lactate, and L-alanine are the major organic end products. Thermotagae fermentation processes are influenced by various factors, such as hydrogen partial pressure, agitation, gas sparging, culture/headspace ratio, inoculum, pH, temperature, nitrogen sources, sulfur sources, inorganic compounds, metal ions, etc. Optimization of these parameters will help to fully unleash the biotechnological potentials of Thermotogae and promote their applications in industry. This article gives an overview of how these operational parameters could impact Thermotogae fermentation in terms of sugar consumption, hydrogen yields, and organic acids production. Full article
(This article belongs to the Special Issue Thermophilic and Hyperthermophilic Microbes and Enzymes)
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19 pages, 12084 KiB  
Review
O6-alkylguanine-DNA Alkyltransferases in Microbes Living on the Edge: From Stability to Applicability
by Rosanna Mattossovich, Rosa Merlo, Riccardo Miggiano, Anna Valenti and Giuseppe Perugino
Int. J. Mol. Sci. 2020, 21(8), 2878; https://0-doi-org.brum.beds.ac.uk/10.3390/ijms21082878 - 20 Apr 2020
Cited by 8 | Viewed by 3526
Abstract
The genome of living cells is continuously exposed to endogenous and exogenous attacks, and this is particularly amplified at high temperatures. Alkylating agents cause DNA damage, leading to mutations and cell death; for this reason, they also play a central role in chemotherapy [...] Read more.
The genome of living cells is continuously exposed to endogenous and exogenous attacks, and this is particularly amplified at high temperatures. Alkylating agents cause DNA damage, leading to mutations and cell death; for this reason, they also play a central role in chemotherapy treatments. A class of enzymes known as AGTs (alkylguanine-DNA-alkyltransferases) protects the DNA from mutations caused by alkylating agents, in particular in the recognition and repair of alkylated guanines in O6-position. The peculiar irreversible self-alkylation reaction of these enzymes triggered numerous studies, especially on the human homologue, in order to identify effective inhibitors in the fight against cancer. In modern biotechnology, engineered variants of AGTs are developed to be used as protein tags for the attachment of chemical ligands. In the last decade, research on AGTs from (hyper)thermophilic sources proved useful as a model system to clarify numerous phenomena, also common for mesophilic enzymes. This review traces recent progress in this class of thermozymes, emphasizing their usefulness in basic research and their consequent advantages for in vivo and in vitro biotechnological applications. Full article
(This article belongs to the Special Issue Thermophilic and Hyperthermophilic Microbes and Enzymes)
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