scholarly journals The thermophilic biomass-degrading bacterium Caldicellulosiruptor bescii utilizes two enzymes to oxidize glyceraldehyde 3-phosphate during glycolysis

2019 ◽  
Vol 294 (25) ◽  
pp. 9995-10005 ◽  
Author(s):  
Israel M. Scott ◽  
Gabriel M. Rubinstein ◽  
Farris L. Poole ◽  
Gina L. Lipscomb ◽  
Gerrit J. Schut ◽  
...  
Keyword(s):  
Large Subunit ◽  
Small Subunit ◽  
Lag Phase ◽  
Cellulolytic Bacterium ◽  
Ferredoxin Oxidoreductase ◽  
Caldicellulosiruptor Bescii ◽  
Degrading Bacterium ◽  
Growth Optimum ◽  
Cell Densities ◽  
Aldehyde Ferredoxin Oxidoreductase

Caldicellulosiruptor bescii is an extremely thermophilic, cellulolytic bacterium with a growth optimum at 78 °C and is the most thermophilic cellulose degrader known. It is an attractive target for biotechnological applications, but metabolic engineering will require an in-depth understanding of its primary pathways. A previous analysis of its genome uncovered evidence that C. bescii may have a completely uncharacterized aspect to its redox metabolism, involving a tungsten-containing oxidoreductase of unknown function. Herein, we purified and characterized this new member of the aldehyde ferredoxin oxidoreductase family of tungstoenzymes. We show that it is a heterodimeric glyceraldehyde-3-phosphate (GAP) ferredoxin oxidoreductase (GOR) present not only in all known Caldicellulosiruptor species, but also in 44 mostly anaerobic bacterial genera. GOR is phylogenetically distinct from the monomeric GAP-oxidizing enzyme found previously in several Archaea. We found that its large subunit (GOR-L) contains a single tungstopterin site and one iron-sulfur [4Fe-4S] cluster, that the small subunit (GOR-S) contains four [4Fe-4S] clusters, and that GOR uses ferredoxin as an electron acceptor. Deletion of either subunit resulted in a distinct growth phenotype on both C5 and C6 sugars, with an increased lag phase, but higher cell densities. Using metabolomics and kinetic analyses, we show that GOR functions in parallel with the conventional GAP dehydrogenase, providing an alternative ferredoxin-dependent glycolytic pathway. These two pathways likely facilitate the recycling of reduced redox carriers (NADH and ferredoxin) in response to environmental H2 concentrations. This metabolic flexibility has important implications for the future engineering of this and related species.

scholarly journals Molecular Analysis of a Novel Methanesulfonic Acid Monooxygenase from the Methylotroph Methylosulfonomonas methylovora

1999 ◽  
Vol 181 (7) ◽  
pp. 2244-2251 ◽  
Author(s):  
Paolo de Marco ◽  
Pedro Moradas-Ferreira ◽  
Timothy P. Higgins ◽  
Ian McDonald ◽  
Elizabeth M. Kenna ◽  
...  
Keyword(s):  
Large Subunit ◽  
Methanesulfonic Acid ◽  
Small Subunit ◽  
Methylotrophic Bacterium ◽  
Binding Motif ◽  
Chromosomal Dna ◽  
Mononuclear Iron ◽  
Degrading Bacterium ◽  
Genes Encoding ◽  
High Degree

ABSTRACT Methylosulfonomonas methylovora M2 is an unusual gram-negative methylotrophic bacterium that can grow on methanesulfonic acid (MSA) as the sole source of carbon and energy. Oxidation of MSA by this bacterium is carried out by a multicomponent MSA monooxygenase (MSAMO). Cloning and sequencing of a 7.5-kbp SphI fragment of chromosomal DNA revealed four tightly linked genes encoding this novel monooxygenase. Analysis of the deduced MSAMO polypeptide sequences indicated that the enzyme contains a two-component hydroxylase of the mononuclear-iron-center type. The large subunit of the hydroxylase, MsmA (48 kDa), contains a typical Rieske-type [2Fe–2S] center with an unusual iron-binding motif and, together with the small subunit of the hydroxylase, MsmB (20 kDa), showed a high degree of identity with a number of dioxygenase enzymes. However, the other components of the MSAMO, MsmC, the ferredoxin component, and MsmD, the reductase, more closely resemble those found in other classes of oxygenases. MsmC has a high degree of identity to ferredoxins from toluene and methane monooxygenases, which are enzymes characterized by possessing hydroxylases containing μ-oxo bridge binuclear iron centers. MsmD is a reductase of 38 kDa with a typical chloroplast-like [2Fe–2S] center and conserved flavin adenine dinucleotide- and NAD-binding motifs and is similar to a number of mono- and dioxygenase reductase components. Preliminary analysis of the genes encoding MSAMO from a marine MSA-degrading bacterium, Marinosulfonomonas methylotropha, revealed the presence of msm genes highly related to those found in Methylosulfonomonas, suggesting that MSAMO is a novel type of oxygenase that may be conserved in all MSA-utilizing bacteria.

Depletion and Reconstitution of Active E. Coli Ribosomes

1975 ◽  
Vol 33 ◽  
pp. 640-641
Author(s):  
M. Boublik ◽  
W. Hellmann ◽  
F. Jenkins
Keyword(s):  
Protein Biosynthesis ◽  
Large Subunit ◽  
High Resolution Electron Microscopy ◽  
Small Subunit ◽  
Structure And Function ◽  
Biologically Active ◽  
Biological Macromolecules ◽  
E Coli ◽  
Resolution Electron ◽  
And Function

Correlations between structure and function of biological macromolecules have been studied intensively for many years, mostly by indirect methods. High resolution electron microscopy is a unique tool which can provide such information directly by comparing the conformation of biopolymers in their biologically active and inactive state. We have correlated the structure and function of ribosomes, ribonucleoprotein particles which are the site of protein biosynthesis. 70S E. coli ribosomes, used in this experiment, are composed of two subunits - large (50S) and small (30S). The large subunit consists of 34 proteins and two different ribonucleic acid molecules. The small subunit contains 21 proteins and one RNA molecule. All proteins (with the exception of L7 and L12) are present in one copy per ribosome.This study deals with the changes in the fine structure of E. coli ribosomes depleted of proteins L7 and L12. These proteins are unique in many aspects.

Theileria parva ribosomal internal transcribed spacer sequences exhibit extensive polymorphism and mosaic evolution: application to the characterization of parasites from cattle and buffalo

Parasitology ◽  
1999 ◽  
Vol 118 (6) ◽  
pp. 541-551 ◽  
Author(s):  
N. E. COLLINS ◽  
B. A. ALLSOPP
Keyword(s):  
Genetic Recombination ◽  
Large Subunit ◽  
Small Subunit ◽  
Internal Transcribed Spacers ◽  
Rrna Genes ◽  
Gene Pools ◽  
Theileria Parva ◽  
Causative Agents ◽  
Internal Transcribed Spacer Sequences

We sequenced the rRNA genes and internal transcribed spacers (ITS) of several Theileria parva isolates in an attempt to distinguish between the causative agents of East coast fever and Corridor disease. The small subunit (SSU) and large subunit (LSU) rRNA genes from a cloned T. p. lawrencei parasite were sequenced; the former was identical to that of T. p. parva Muguga, and there were minor heterogeneities in the latter. The 5·8S gene sequences of 11 T. parva isolates were identical, but major differences were found in the ITS. Six characterization oligonucleotides were designed to hybridize within the variable ITS1 region; 93·5% of T. p. parva isolates examined were detected by probe TPP1 and 81·8% of T. p. lawrencei isolates were detected by TPL2 and/or TPL3a. There was no absolute distinction between T. p. parva and T. p. lawrencei and the former hybridized with fewer of the probes than did the latter. It therefore seems that a relatively homogenous subpopulation of T. parva has been selected in cattle from a more diverse gene pool in buffalo. The ITSs of both T. p. parva and T. p. lawrencei contained different combinations of identifiable sequence segments, resulting in a mosaic of segments in any one isolate, suggesting that the two populations undergo genetic recombination and that their gene pools are not completely separate.

scholarly journals Domains for protein-protein interactions at the N and C termini of the large subunit of bacteriophage lambda terminase.

Genetics ◽  
1988 ◽  
Vol 119 (3) ◽  
pp. 477-484
Author(s):  
W F Wu ◽  
S Christiansen ◽  
M Feiss
Keyword(s):  
Protein Interactions ◽  
Ternary Complex ◽  
Large Subunit ◽  
Small Subunit ◽  
Functional Domain ◽  
Binary Complex ◽  
Protein Protein Interactions ◽  
Related Phage ◽  
Carboxy Terminal ◽  
Lambda Dna

Abstract The large subunit of phage lambda terminase, gpA, the gene product of the phage A gene, interacts with the small subunit, gpNul, to form functional terminase. Terminase binds to lambda DNA at cosB to form a binary complex. The terminase:DNA complex binds a prohead to form a ternary complex. Ternary complex formation involves an interaction of the prohead with gpA. The amino terminus of gpA contains a functional domain for interaction with gpNul, and the carboxy-terminal 38 amino acids of gpA contain a functional domain for prohead binding. This information about the structure of gpA was obtained through the use of hybrid phages resulting from recombination between lambda and the related phage 21. lambda and 21 encode terminases that are analogous in structural organization and have ca. 60% sequence identity. In spite of these similarities, lambda and 21 terminases differ in specificity for DNA binding, subunit assembly, and prohead binding. A lambda-21 hybrid phage produces a terminase in which one of the subunits is chimeric and had recombinant specificities. In the work reported here; a new hybrid, lambda-21 hybrid 67, is characterized. lambda-21 hybrid 67 is the result of a crossover between lambda and 21 in the large subunit genes, such that the DNA from the left chromosome end is from 21, including cosB phi 21, the 1 gene, and the first 48 codons for the 2 gene. The rest of the hybrid 67 chromosome is lambda DNA, including 593 codons of the A gene. The chimeric gp2/A of hybrid 67 binds gp1 to form functional terminase.(ABSTRACT TRUNCATED AT 250 WORDS)

scholarly journals The Evolution of Life Modes in Stictidaceae, with Three Novel Taxa

2021 ◽  
Vol 7 (2) ◽  
pp. 105
Author(s):  
Vinodhini Thiyagaraja ◽  
Robert Lücking ◽  
Damien Ertz ◽  
Samantha C. Karunarathna ◽  
Dhanushka N. Wanasinghe ◽  
...  
Keyword(s):  
New Species ◽  
Large Scale ◽  
Sequence Data ◽  
Large Subunit ◽  
Monte Carlo Sampling ◽  
Small Subunit ◽  
Sensu Stricto ◽  
Internal Transcribed Spacers ◽  
Character State ◽  
Phenotypic Data

Ostropales sensu lato is a large group comprising both lichenized and non-lichenized fungi, with several lineages expressing optional lichenization where individuals of the same fungal species exhibit either saprotrophic or lichenized lifestyles depending on the substrate (bark or wood). Greatly variable phenotypic characteristics and large-scale phylogenies have led to frequent changes in the taxonomic circumscription of this order. Ostropales sensu lato is currently split into Graphidales, Gyalectales, Odontotrematales, Ostropales sensu stricto, and Thelenellales. Ostropales sensu stricto is now confined to the family Stictidaceae, which includes a large number of species that are poorly known, since they usually have small fruiting bodies that are rarely collected, and thus, their taxonomy remains partly unresolved. Here, we introduce a new genus Ostropomyces to accommodate a novel lineage related to Ostropa, which is composed of two new species, as well as a new species of Sphaeropezia, S. shangrilaensis. Maximum likelihood and Bayesian inference analyses of mitochondrial small subunit spacers (mtSSU), large subunit nuclear rDNA (LSU), and internal transcribed spacers (ITS) sequence data, together with phenotypic data documented by detailed morphological and anatomical analyses, support the taxonomic affinity of the new taxa in Stictidaceae. Ancestral character state analysis did not resolve the ancestral nutritional status of Stictidaceae with confidence using Bayes traits, but a saprotrophic ancestor was indicated as most likely in a Bayesian binary Markov Chain Monte Carlo sampling (MCMC) approach. Frequent switching in nutritional modes between lineages suggests that lifestyle transition played an important role in the evolution of this family.

Morphometrics and molecular analysis ofOzolaimus linstowin. sp. (Oxyuroidea: Pharyngodonidae) from the green lizardIguana iguana

2015 ◽  
Vol 90 (2) ◽  
pp. 186-198 ◽  
Author(s):  
S.V. Malysheva
Keyword(s):  
Body Size ◽  
Large Subunit ◽  
Buccal Capsule ◽  
Small Subunit ◽  
Present Species ◽  
Lsu Rdna ◽  
Adult Males ◽  
Spicule Length ◽  
Males And Females ◽  
Characteristic Features

AbstractOzolaimus linstowin. sp. is described from the large intestine ofIguana iguanaLinnaeus, 1758 from Mexico. The present species can be easily distinguished fromO. megatyphlonandO. cirratusby the presence of a long and slender pharynx not divided into sections, more similar to the remaining two species,O. monhysteraandO. ctenosauri. Ozolaimus linstowin. sp. can be differentiated fromO. monhysteraby the shorter spicule length and smaller body size of both males and females. Males ofO. linstowin. sp. are morphologically close to those ofO. ctenosauri, but females possess a markedly smaller body size and differ in the organization of the oral cuticular armature. Adult males ofO. linstowin. sp. bear some characteristic features of the J3 juvenile morphology in terms of the cuticular organization of the oral and buccal capsule. Phylogenetic analysis ofO.linstowin. sp. using partial small subunit (SSU) and D2–D3 large subunit (LSU) rDNA shows relationships with several Oxyuridae genera.

scholarly journals Molecular Diversity of the Syndinean Genus Euduboscquella Based on Single-Cell PCR Analysis

2011 ◽  
Vol 78 (2) ◽  
pp. 334-345 ◽  
Author(s):  
Tsvetan R. Bachvaroff ◽  
Sunju Kim ◽  
Laure Guillou ◽  
Charles F. Delwiche ◽  
D. Wayne Coats
Keyword(s):  
Large Subunit ◽  
Small Subunit ◽  
Base Change ◽  
Pcr Analysis ◽  
Ssu Rrna ◽  
Compensatory Base Change ◽  
Content Type ◽  
Single Cell Pcr ◽  
Group I ◽  
Lsu Rrna

ABSTRACTThe genusEuduboscquellais one of a few described genera within the syndinean dinoflagellates, an enigmatic lineage with abundant diversity in marine environmental clone libraries based on small subunit (SSU) rRNA. The region composed of the SSU through to the partial large subunit (LSU) rRNA was determined from 40 individual tintinnid ciliate loricae infected withEuduboscquellasampled from eight surface water sites in the Northern Hemisphere, producing seven distinct SSU sequences. The corresponding host SSU rRNA region was also amplified from eight host species. The SSU tree ofEuduboscquellaand syndinean group I sequences from environmental clones had seven well-supported clades and one poorly supported clade across data sets from 57 to 692 total sequences. The genusEuduboscquellaconsistently formed a supported monophyletic clade within a single subclade of group I sequences. For most parasites with identical SSU sequences, the more variable internal transcribed spacer (ITS) to LSU rRNA regions were polymorphic at 3 to 10 sites. However, inE. cachonithere was variation between ITS to LSU copies at up to 20 sites within an individual, while in a parasite ofTintinnopsisspp., variation between different individuals ranged up to 19 polymorphic sites. However, applying the compensatory base change model to the ITS2 sequences suggested no compensatory changes within or between individuals with the same SSU sequence, while one to four compensatory changes between individuals with similar but not identical SSU sequences were found. Comparisons between host and parasite phylogenies do not suggest a simple pattern of host or parasite specificity.

scholarly journals Investigation of in Vivo Roles of the C-terminal Tails of the Small Subunit (ββ′) of Saccharomyces cerevisiae Ribonucleotide Reductase

2013 ◽  
Vol 288 (20) ◽  
pp. 13951-13959 ◽  
Author(s):  
Yan Zhang ◽  
Xiuxiang An ◽  
JoAnne Stubbe ◽  
Mingxia Huang
Keyword(s):  
Saccharomyces Cerevisiae ◽  
Ribonucleotide Reductase ◽  
Large Subunit ◽  
Small Subunit ◽  
Cluster Assembly ◽  
E Coli ◽  
Viability Assay ◽  
Docking Model

The small subunit (β2) of class Ia ribonucleotide reductase (RNR) houses a diferric tyrosyl cofactor (Fe2III-Y•) that initiates nucleotide reduction in the large subunit (α2) via a long range radical transfer (RT) pathway in the holo-(α2)m(β2)n complex. The C-terminal tails of β2 are predominantly responsible for interaction with α2, with a conserved tyrosine residue in the tail (Tyr356 in Escherichia coli NrdB) proposed to participate in cofactor assembly/maintenance and in RT. In the absence of structure of any holo-RNR, the role of the β tail in cluster assembly/maintenance and its predisposition within the holo-complex have remained unknown. In this study, we have taken advantage of the unusual heterodimeric nature of the Saccharomyces cerevisiae RNR small subunit (ββ′), of which only β contains a cofactor, to address both of these issues. We demonstrate that neither β-Tyr376 nor β′-Tyr323 (Tyr356 equivalent in NrdB) is required for cofactor assembly in vivo, in contrast to the previously proposed mechanism for E. coli cofactor maintenance and assembly in vitro. Furthermore, studies with reconstituted-ββ′ and an in vivo viability assay show that β-Tyr376 is essential for RT, whereas Tyr323 in β′ is not. Although the C-terminal tail of β′ is dispensable for cofactor formation and RT, it is essential for interactions with β and α to form the active holo-RNR. Together the results provide the first evidence of a directed orientation of the β and β′ C-terminal tails relative to α within the holoenzyme consistent with a docking model of the two subunits and argue against RT across the β β′ interface.

Morphological and molecular characteristics of Bastiania sinensis sp. n., Tripyla aquatica Brzeski & Winiszewska-Ślipińska, 1993 and T. setifera Bütschli, 1873 (Nematoda: Triplonchida) from Shanxi Province, North China

Nematology ◽  
2021 ◽  
pp. 1-23
Author(s):  
Mei Na Liu ◽  
Yu Mei Xu ◽  
Zeng Qi Zhao ◽  
Jian Ming Wang
Keyword(s):  
North China ◽  
Large Subunit ◽  
Small Subunit ◽  
Rrna Genes ◽  
Shanxi Province ◽  
Molecular Characteristics ◽  
Separate Species ◽  
Using Data ◽  
First Time ◽  
Respective Species

Summary This paper describes a new species of Bastiania, presents a new record and redescribes a known species of Tripyla. These nematodes are all in the order Triplonchida and were collected from Shanxi Province, North China. Bastiania sinensis sp. n. is characterised by having the female with a relatively slender body 1049-1295 μm long, dorsally arcuate after heat relaxation, with outer labial setae and cephalic setae in a single circle, an oval amphid, 7-8 laterodorsal cervical setae scattered in the pharyngeal region, orthometamenes and pseudocoelomocytes present, tail conoid with a mucron 1-2 μm long, two pairs of caudal setae present, a = 58.1-75.5, b = 4.0-4.6, c = 12.7-19.7, c′ = 4.1-7.8 and V = 61.1-67.7. Males were not found. Tripyla aquatica is recorded for the first time from China, and is redescribed. Tripyla setifera has been reported from China but without a detailed description – now provided. In addition, phylogenetic relationships among the species were analysed using data from the near full length small subunit (SSU) and D2-D3 segments of large subunit (LSU) of rRNA genes. Bastiania sinensis sp. n. is monophyletic with the Bastiania sequences available in GenBank, but is on an independent branch supporting its status as a separate species; T. aquatica and T. setifera are monophyletically clustered with known Tripyla species and grouped together with sequences from their respective species.

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