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Search indexed NASA NTRS and DOE OSTI research on propulsion, heat transfer, battery materials and energy systems. Follow report and document links to the original sources.

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At least 631 records · Page 35
All sourcesNASA NTRSDOE OSTI
DOE OSTI2015

period -1 encodes an ATP-dependent RNA helicase that influences nutritional compensation of the Neurospora circadian clock

Mutants in the period -1 ( prd­ -1) gene, characterized by a recessive allele, display a reduced growth rate and period lengthening of the developmental cycle controlled by the circadian clock. We refined the genetic location of prd­ -1 and used whole genome sequencing to find the mutation defining it, confirming the identity of prd­ -1 by rescuing the mutant circadian phenotype via transformation. PRD-1 is an RNA helicase whose orthologs, DDX5 and DDX17 in humans and Dbp2p in yeast, are implicated in various processes including transcriptional regulation, elongation, and termination, 23 ribosome biogenesis, and RNA decay. Although prd­ -1smutantssiois an ATP-dependent RNA helicase, member of a sub-family display a long period (~25 hrs) circadian developmental cycle, they interestingly display a wild type period when the core circadian oscillator is tracked using a frq -luciferase transcriptional fusion under conditions of limiting nutritional carbon; the core oscillator runs with a long period under glucose-sufficient conditions. Furthermore PRD-1 clearly impacts the circadian oscillator and is not only part of a metabolic oscillator ancillary to the core clock. PRD-1 is an essential protein and its expression is neither light-regulated nor clock-regulated. However, it is transiently induced by glucose; in the presence of sufficient glucose PRD-1 is in the nucleus until glucose runs out which elicits its disappearance from the nucleus. Because circadian period length is carbon concentration-dependent, prd­ -1 may be formally viewed as clock mutant with defective nutritional compensation of circadian period length.

59 BASIC BIOLOGICAL SCIENCES↗
DOE OSTI2016

Methanobactin transport machinery

Methanotrophic bacteria use methane, a potent greenhouse gas, as their primary source of carbon and energy. The first step in methane metabolism is its oxidation to methanol. In almost all methanotrophs, this chemically challenging reaction is catalyzed by particulate methane monooxygenase (pMMO), a copper-dependent integral membrane enzyme. Methanotrophs acquire copper (Cu) for pMMO by secreting a small ribosomally produced, posttranslationally modified natural product called methanobactin (Mbn). Mbn chelates Cu with high affinity, and the Cu-loaded form (CuMbn) is reinternalized into the cell via an active transport process. Bioinformatic and gene regulation studies suggest that two proteins might play a role in CuMbn handling: the TonB-dependent transporter MbnT and the periplasmic binding protein MbnE. Disruption of the gene that encodes MbnT abolishes CuMbn uptake, as reported previously, and expression of MbnT in Escherichia coli confers the ability to take up CuMbn. Biophysical studies of MbnT and MbnE reveal specific interactions with CuMbn, and a crystal structure of apo MbnE is consistent with MbnE's proposed role as a periplasmic CuMbn transporter. Notably, MbnT and MbnE exhibit different levels of discrimination between cognate and noncognate CuMbns. Furthermore, these findings provide evidence for CuMbn–protein interactions and begin to elucidate the molecular mechanisms of its recognition and transport.

59 BASIC BIOLOGICAL SCIENCES↗
DOE OSTI2016

Emergent rules for codon choice elucidated by editing rare arginine codons in Escherichia coli

Significance This work presents the genome-wide replacement of all rare AGR (AGA and AGG) arginine codons in the essential genes of Escherichia coli with synonymous CGN alternatives. Synonymous codon substitutions can lethally impact noncoding function by disrupting mRNA secondary structure and ribosomal binding site-like motifs. Here we quantitatively define the range of tolerable deviation in these metrics and use this relationship to provide critical insight into codon choice in recoded genomes. This work demonstrates that genome-wide removal of AGR is likely to be possible and provides a framework for designing genomes with radically altered genetic codes.

59 BASIC BIOLOGICAL SCIENCES↗
DOE OSTI2017

Foldamer hypothesis for the growth and sequence differentiation of prebiotic polymers

It is not known how life originated. It is thought that prebiotic processes were able to synthesize short random polymers. However, then, how do short-chain molecules spontaneously grow longer? Also, how would random chains grow more informational and become autocatalytic (i.e., increasing their own concentrations)? We study the folding and binding of random sequences of hydrophobic ( H ) and polar ( P ) monomers in a computational model. We find that even short hydrophobic polar ( HP ) chains can collapse into relatively compact structures, exposing hydrophobic surfaces. In this way, they act as primitive versions of today’s protein catalysts, elongating other such HP polymers as ribosomes would now do. Such foldamer catalysts are shown to form an autocatalytic set, through which short chains grow into longer chains that have particular sequences. An attractive feature of this model is that it does not overconverge to a single solution; it gives ensembles that could further evolve under selection. This mechanism describes how specific sequences and conformations could contribute to the chemistry-to-biology (CTB) transition.

36 MATERIALS SCIENCE↗
DOE OSTI2017

Characterization of the macrocyclase involved in the biosynthesis of RiPP cyclic peptides in plants

Enzymes that can catalyze the macrocyclization of linear peptide substrates have long been sought for the production of libraries of structurally diverse scaffolds via combinatorial gene assembly as well as to afford rapid in vivo screening methods. Orbitides are plant ribosomally synthesized and posttranslationally modified peptides (RiPPs) of various sizes and topologies, several of which are shown to be biologically active. The diversity in size and sequence of orbitides suggests that the corresponding macrocyclases may be ideal catalysts for production of cyclic peptides. Here we present the biochemical characterization and crystal structures of the plant enzyme PCY1 involved in orbitide macrocyclization. These studies demonstrate how the PCY1 S9A protease fold has been adapted for transamidation, rather than hydrolysis, of acyl-enzyme intermediates to yield cyclic products. Notably, PCY1 uses an unusual strategy in which the cleaved C-terminal follower peptide from the substrate stabilizes the enzyme in a productive conformation to facilitate macrocyclization of the N-terminal fragment. The broad substrate tolerance of PCY1 can be exploited as a biotechnological tool to generate structurally diverse arrays of macrocycles, including those with nonproteinogenic elements.

59 BASIC BIOLOGICAL SCIENCES↗
DOE OSTI2017

Structural insights into enzymatic [4+2] aza -cycloaddition in thiopeptide antibiotic biosynthesis

The [4+2] cycloaddition reaction is an enabling transformation in modern synthetic organic chemistry, but there are only limited examples of dedicated natural enzymes that can catalyze this transformation. Thiopeptides (or more formally thiazolyl peptides) are a class of thiazole-containing, highly modified, macrocyclic secondary metabolites made from ribosomally synthesized precursor peptides. The characteristic feature of these natural products is a six-membered nitrogenous heterocycle that is assembled via a formal [4+2] cycloaddition between two dehydroalanine (Dha) residues. This heteroannulation is entirely contingent on enzyme activity, although the mechanism of the requisite pyridine/dehydropiperidine synthase remains to be elucidated. The unusual aza -cylic product is distinct from the more common carbocyclic products of synthetic and biosynthetic [4+2] cycloaddition reactions. Here, to elucidate the mechanism of cycloaddition, we have determined atomic resolution structures of the pyridine synthases involved in the biosynthesis of the thiopeptides thiomuracin (TbtD) and GE2270A (PbtD), in complex with substrates and product analogs. Structure-guided biochemical, mutational, computational, and binding studies elucidate active-site features that explain how orthologs can generate rigid macrocyclic scaffolds of different sizes. Notably, the pyridine synthases show structural similarity to the elimination domain of lanthipeptide dehydratases, wherein insertions of secondary structural elements result in the formation of a distinct active site that catalyzes different chemistry. Comparative analysis identifies other catalysts that contain a shared core protein fold but whose active sites are located in entirely different regions, illustrating a principle predicted from efforts in de novo protein design.

59 BASIC BIOLOGICAL SCIENCES↗
DOE OSTI2019

Direct visualization of degradation microcompartments at the ER membrane

To promote the biochemical reactions of life, cells can compartmentalize molecular interaction partners together within separated non–membrane-bound regions. It is unknown whether this strategy is used to facilitate protein degradation at specific locations within the cell. Leveraging in situ cryo-electron tomography to image the native molecular landscape of the unicellular alga Chlamydomonas reinhardtii , we discovered that the cytosolic protein degradation machinery is concentrated within ∼200-nm foci that contact specialized patches of endoplasmic reticulum (ER) membrane away from the ER–Golgi interface. These non–membrane-bound microcompartments exclude ribosomes and consist of a core of densely clustered 26S proteasomes surrounded by a loose cloud of Cdc48. Active proteasomes in the microcompartments directly engage with putative substrate at the ER membrane, a function canonically assigned to Cdc48. Live-cell fluorescence microscopy revealed that the proteasome clusters are dynamic, with frequent assembly and fusion events. We propose that the microcompartments perform ER-associated degradation, colocalizing the degradation machinery at specific ER hot spots to enable efficient protein quality control.

Science & Technology - Other Topics↗
DOE OSTI2019

Steric complementarity directs sequence promiscuous leader binding in RiPP biosynthesis

Enzymes that generate ribosomally synthesized and posttranslationally modified peptide (RiPP) natural products have garnered significant interest, given their ability to produce large libraries of chemically diverse scaffolds. Such RiPP biosynthetic enzymes are predicted to bind their corresponding peptide substrates through sequence-specific recognition of the leader sequence, which is removed after the installation of posttranslational modifications on the core sequence. The conservation of the leader sequence within a given RiPP class, in otherwise disparate precursor peptides, further supports the notion that strict sequence specificity is necessary for leader peptide engagement. Here, we demonstrate that leader binding by a biosynthetic enzyme in the lasso peptide class of RiPPs is directed by a minimal number of hydrophobic interactions. Biochemical and structural data illustrate how a single leader-binding domain can engage sequence-divergent leader peptides using a conserved motif that facilitates hydrophobic packing. We report the presence of this simple motif in noncognate peptides results in low micromolar affinity binding by binding domains from several different lasso biosynthetic systems. We also demonstrate that these observations likely extend to other RiPP biosynthetic classes. The portability of the binding motif opens avenues for the engineering of semisynthetic hybrid RiPP products.

37 INORGANIC, ORGANIC, PHYSICAL, AND ANALYTICAL CH↗
DOE OSTI2020

A majority of Rhodobacter sphaeroides promoters lack a crucial RNA polymerase recognition feature, enabling coordinated transcription activation

Using an in vitro transcription system with purified RNA polymerase (RNAP) to investigate rRNA synthesis in the photoheterotrophic α-proteobacterium Rhodobacter sphaeroides, we identified a surprising feature of promoters recognized by the major holoenzyme. Transcription from R. sphaeroides rRNA promoters was unexpectedly weak, correlating with absence of 7T, the very highly conserved thymine found at the last position in –10 elements of promoters in most bacterial species. Thymine substitutions for adenine at position –7 in the three rRNA promoters strongly increased intrinsic promoter activity, indicating that R. sphaeroides RNAP can utilize –7T when present. rRNA promoters were activated by purified R. sphaeroides CarD, a transcription factor found in many bacterial species but not in β- and γ-proteobacteria. Overall, CarD increased the activity of 15 of 16 native R. sphaeroides promoters tested in vitro that lacked –7T, whereas it had no effect on three of the four native promoters that contained –7T. Genome-wide bioinformatic analysis of promoters from R. sphaeroides and two other α-proteobacterial species indicated that 30 to 43% contained –7T, whereas 90 to 99% of promoters from non–α-proteobacteria contained –7T. Thus, promoters lacking –7T appear to be widespread in α-proteobacteria and may have evolved away from consensus to enable their coordinated regulation by transcription factors like CarD. We observed a strong reduction in R. sphaeroides CarD levels when cells enter stationary phase, suggesting that reduced activation by CarD may contribute to inhibition of rRNA transcription when cells enter stationary phase, the stage of growth when bacterial ribosome synthesis declines.

59 BASIC BIOLOGICAL SCIENCES↗
DOE OSTI2023

ADP enhances the allosteric activation of eukaryotic elongation factor 2 kinase by calmodulin

Protein translation, one of the most energy-consumptive processes in a eukaryotic cell, requires robust regulation, especially under energy-deprived conditions. A critical component of this regulation is the suppression of translational elongation through reduced ribosome association of the GTPase eukaryotic elongation factor 2 (eEF-2) resulting from its specific phosphorylation by the calmodulin (CaM)-activated α–kinase eEF-2 kinase (eEF-2K). It has been suggested that the eEF-2K response to reduced cellular energy levels is indirect and mediated by the universal energy sensor AMP-activated protein kinase (AMPK) through direct stimulatory phosphorylation and/or downregulation of the eEF-2K-inhibitory nutrient-sensing mTOR pathway. Here, we provide structural, biochemical, and cell-biological evidence of a direct energy-sensing role of eEF-2K through its stimulation by ADP. A crystal structure of the nucleotide-bound complex between CaM and the functional core of eEF-2K phosphorylated at its primary stimulatory site (T348) reveals ADP bound at a unique pocket located on the face opposite that housing the kinase active site. Within this basic pocket (BP), created at the CaM/eEF-2K interface upon complex formation, ADP is stabilized through numerous interactions with both interacting partners. Biochemical analyses using wild-type eEF-2K and specific BP mutants indicate that ADP stabilizes CaM within the active complex, increasing the sensitivity of the kinase to CaM. Induction of energy stress through glycolysis inhibition results in significantly reduced enhancement of phosphorylated eEF-2 levels in cells expressing ADP-binding compromised BP mutants compared to cells expressing wild-type eEF-2K. These results suggest a direct energy-sensing role for eEF-2K through its cooperative interaction with CaM and ADP.

59 BASIC BIOLOGICAL SCIENCES↗
DOE OSTI2024

Development of 2nd generation aminomethyl spectinomycins that overcome native efflux in Mycobacterium abscessus

Mycobacterium abscessus (Mab), a nontuberculous mycobacterial (NTM) species, is an emerging pathogen with high intrinsic drug resistance. Current standard-of-care therapy results in poor outcomes, demonstrating the urgent need to develop effective antimycobacterial regimens. Through synthetic modification of spectinomycin (SPC), we have identified a distinct structural subclass of N-ethylene linked aminomethyl SPCs (eAmSPCs) that are up to 64-fold more potent against Mab over the parent SPC. Mechanism of action and crystallography studies demonstrate that the eAmSPCs display a mode of ribosomal inhibition consistent with SPC. However, they exert their increased antimicrobial activity through enhanced accumulation, largely by circumventing efflux mechanisms. The N-ethylene linkage within this series plays a critical role in avoiding TetV-mediated efflux, as lead eAmSPC 2593 displays a mere fourfold susceptibility improvement against Mab ΔtetV, in contrast to the 64-fold increase for SPC. Even a minor shortening of the linkage by a single carbon, akin to 1st generation AmSPC 1950, results in a substantial increase in MICs and a 16-fold rise in susceptibility against Mab ΔtetV. These shifts suggest that longer linkages might modify the kinetics of drug expulsion by TetV, ultimately shifting the equilibrium towards heightened intracellular concentrations and enhanced antimicrobial efficacy. Furthermore, lead eAmSPCs were also shown to synergize with various classes of anti-Mab antibiotics and retain activity against clinical isolates and other mycobacterial strains. Encouraging pharmacokinetic profiles coupled with robust efficacy in Mab murine infection models suggest that eAmSPCs hold the potential to be developed into treatments for Mab and other NTM infections.

59 BASIC BIOLOGICAL SCIENCES↗
DOE OSTI2024

An essential and highly selective protein import pathway encoded by nucleus-forming phage

Targeting proteins to specific subcellular destinations is essential in prokaryotes, eukaryotes, and the viruses that infect them. Chimalliviridae phages encapsulate their genomes in a nucleus-like replication compartment composed of the protein chimallin (ChmA) that excludes ribosomes and decouples transcription from translation. These phages selectively partition proteins between the phage nucleus and the bacterial cytoplasm. Currently, the genes and signals that govern selective protein import into the phage nucleus are unknown. Here, we identify two components of this protein import pathway: a species-specific surface-exposed region of a phage intranuclear protein required for nuclear entry and a conserved protein, PicA (Protein importer of chimalliviruses A), that facilitates cargo protein trafficking across the phage nuclear shell. We also identify a defective cargo protein that is targeted to PicA on the nuclear periphery but fails to enter the nucleus, providing insight into the mechanism of nuclear protein trafficking. Using CRISPRi-ART protein expression knockdown of PicA, we show that PicA is essential early in the chimallivirus replication cycle. Together, our results allow us to propose a multistep model for the Protein Import Chimallivirus pathway, where proteins are targeted to PicA by amino acids on their surface and then licensed by PicA for nuclear entry. The divergence in the selectivity of this pathway between closely related chimalliviruses implicates its role as a key player in the evolutionary arms race between competing phages and their hosts.

59 BASIC BIOLOGICAL SCIENCES↗
DOE OSTI2024

A potential role for RNA aminoacylation prior to its role in peptide synthesis

Coded ribosomal peptide synthesis could not have evolved unless its sequence and amino acid–specific aminoacylated tRNA substrates already existed. We therefore wondered whether aminoacylated RNAs might have served some primordial function prior to their role in protein synthesis. Here, we show that specific RNA sequences can be nonenzymatically aminoacylated and ligated to produce amino acid–bridged stem-loop RNAs. We used deep sequencing to identify RNAs that undergo highly efficient glycine aminoacylation followed by loop-closing ligation. The crystal structure of one such glycine-bridged RNA hairpin reveals a compact internally stabilized structure with the same eponymous T-loop architecture that is found in many noncoding RNAs, including the modern tRNA. We demonstrate that the T-loop-assisted amino acid bridging of RNA oligonucleotides enables the rapid template-free assembly of a chimeric version of an aminoacyl-RNA synthetase ribozyme. We suggest that the primordial assembly of amino acid–bridged chimeric ribozymes provides a direct and facile route for the covalent incorporation of amino acids into RNA. A greater functionality of covalently incorporated amino acids could contribute to enhanced ribozyme catalysis, providing a driving force for the evolution of sequence and amino acid–specific aminoacyl-RNA synthetase ribozymes in the RNA World. The synthesis of specifically aminoacylated RNAs, an unlikely prospect for nonenzymatic reactions but a likely one for ribozymes, could have set the stage for the subsequent evolution of coded protein synthesis.

Science & Technology - Other Topics↗
DOE OSTI2011

Terminal Alkene Formation by the Thioesterase of Curacin A Biosynthesis: Structure of a Decarboxylating Thioesterase

Curacin A is a polyketide synthase (PKS)-non-ribosomal peptide synthetase-derived natural product with potent anticancer properties generated by the marine cyanobacterium Lyngbya majuscula. Type I modular PKS assembly lines typically employ a thioesterase (TE) domain to off-load carboxylic acid or macrolactone products from an adjacent acyl carrier protein (ACP) domain. In a striking departure from this scheme the curacin A PKS employs tandem sulfotransferase and TE domains to form a terminal alkene moiety. Sulfotransferase sulfonation of {beta}-hydroxy-acyl-ACP is followed by TE hydrolysis, decarboxylation, and sulfate elimination (Gu, L., Wang, B., Kulkarni, A., Gehret, J. J., Lloyd, K. R., Gerwick, L., Gerwick, W. H., Wipf, P., Hakansson, K., Smith, J. L., and Sherman, D. H. (2009) J. Am. Chem. Soc. 131, 16033-16035). With low sequence identity to other PKS TEs (<15%), the curacin TE represents a new thioesterase subfamily. The 1.7-{angstrom} curacin TE crystal structure reveals how the familiar {alpha}/{beta}-hydrolase architecture is adapted to specificity for {beta}-sulfated substrates. A Ser-His-Glu catalytic triad is centered in an open active site cleft between the core domain and a lid subdomain. Unlike TEs from other PKSs, the lid is fixed in an open conformation on one side by dimer contacts of a protruding helix and on the other side by an arginine anchor from the lid into the core. Adjacent to the catalytic triad, another arginine residue is positioned to recognize the substrate {beta}-sulfate group. The essential features of the curacin TE are conserved in sequences of five other putative bacterial ACP-ST-TE tridomains. Formation of a sulfate leaving group as a biosynthetic strategy to facilitate acyl chain decarboxylation is of potential value as a route to hydrocarbon biofuels.

59 BASIC BIOLOGICAL SCIENCES↗
DOE OSTI2017

Two alternative binding mechanisms connect the protein translocation Sec71-Sec72 complex with heat shock proteins

The biosynthesis of many eukaryotic proteins requires accurate targeting to and translocation across the endoplasmic reticulum membrane. Post-translational protein translocation in yeast requires both the Sec61 translocation channel, and a complex of four additional proteins: Sec63, Sec62, Sec71, and Sec72. The structure and function of these proteins are largely unknown. This pathway also requires the cytosolic Hsp70 protein Ssa1, but whether Ssa1 associates with the translocation machinery to target protein substrates to the membrane is unclear. Here, we use a combined structural and biochemical approach to explore the role of Sec71-Sec72 subcomplex in post-translational protein translocation. To this end, we report a crystal structure of the Sec71-Sec72 complex, which revealed that Sec72 contains a tetratricopeptide repeat (TPR) domain that is anchored to the endoplasmic reticulum membrane by Sec71. We also determined the crystal structure of this TPR domain with a C-terminal peptide derived from Ssa1, which suggests how Sec72 interacts with full-length Ssa1. Surprisingly, Ssb1, a cytoplasmic Hsp70 that binds ribosome-associated nascent polypeptide chains, also binds to the TPR domain of Sec72, even though it lacks the TPR-binding C-terminal residues of Ssa1. We demonstrate that Ssb1 binds through its ATPase domain to the TPR domain, an interaction that leads to inhibition of nucleotide exchange. Taken together, our results suggest that translocation substrates can be recruited to the Sec71-Sec72 complex either post-translationally through Ssa1 or co-translationally through Ssb1.

37 INORGANIC, ORGANIC, PHYSICAL, AND ANALYTICAL CH↗
DOE OSTI2018

Structural and spectroscopic analyses of the sporulation killing factor biosynthetic enzyme SkfB, a bacterial AdoMet radical sactisynthase

Sactipeptides are a subclass of ribosomally synthesized and post-translationally modified peptides (RiPPs). They contain a unique thioether bond, referred to as a sactionine linkage, between the sulfur atom of a cysteine residue and the α-carbon of an acceptor residue. These linkages are formed via radical chemistry and are essential for the spermicidal, antifungal, and antibacterial properties of sactipeptides. Enzymes that form these linkages, called sactisynthases, are AdoMet radical enzymes in the SPASM/Twitch subgroup whose structures are incompletely characterized. Here, we present the X-ray crystal structure to 1.29-Å resolution and Mössbauer analysis of SkfB, a sactisynthase from Bacillus subtilis involved in making sporulation killing factor (SKF). We found that SkfB is a modular enzyme with an N-terminal peptide-binding domain comprising a RiPP recognition element (RRE), a middle domain that forms a classic AdoMet radical partial (β/α) 6 barrel structure and displays AdoMet bound to the [4Fe-4S] cluster, and a C-terminal region characteristic of the so-called Twitch domain housing an auxiliary iron-sulfur cluster. Notably, both crystallography and Mössbauer analyses suggest that SkfB can bind a [2Fe-2S] cluster at the auxiliary cluster site, which has been observed only once before in a SPASM/Twitch auxiliary cluster site in the structure of another AdoMet radical enzyme, the pyrroloquinoline quinone biosynthesis enzyme PqqE. Taken together, our findings indicate that SkfB from B. subtilis represents a unique enzyme containing several structural features observed in other AdoMet radical enzymes.

37 INORGANIC, ORGANIC, PHYSICAL, AND ANALYTICAL CH↗
DOE OSTI2019

MbnH is a diheme MauG-like protein associated with microbial copper homeostasis

Methanobactins (Mbns) are ribosomally-produced, post-translationally modified peptidic copper-binding natural products produced under conditions of copper limitation. Genes encoding Mbn biosynthetic and transport proteins have been identified in a wide variety of bacteria, indicating a broader role for Mbns in bacterial metal homeostasis. Many of the genes in the Mbn operons have been assigned functions, but two genes usually present, mbnP and mbnH, encode uncharacterized proteins predicted to reside in the periplasm. MbnH belongs to the bacterial diheme cytochrome c peroxidase (bCcP)/MauG protein family, and MbnP contains no domains of known function. Here, we performed a detailed bioinformatic analysis of both proteins and have biochemically characterized MbnH from Methylosinus (Ms.) trichosporium OB3b. We note that the mbnH and mbnP genes typically co-occur and are located proximal to genes associated with microbial copper homeostasis. Our bioinformatics analysis also revealed that the bCcP/MauG family is significantly more diverse than originally appreciated, and that MbnH is most closely related to the MauG subfamily. A 2.6 Å resolution structure of Ms. trichosporium OB3b MbnH combined with spectroscopic data and peroxidase activity assays provided evidence that MbnH indeed more closely resembles MauG than bCcPs, although its redox properties are significantly different from those of MauG. The overall similarity of MbnH to MauG suggests that MbnH could post-translationally modify a macromolecule, such as internalized CuMbn or its uncharacterized partner protein, MbnP. Our results indicate that MbnH is a MauG-like diheme protein that is likely involved in microbial copper homeostasis and represents a new family within the bCcP/MauG superfamily.

37 INORGANIC, ORGANIC, PHYSICAL, AND ANALYTICAL CH↗
DOE OSTI2017

Bifiguratus adelaidae, gen. et sp. nov., a new member of Mucoromycotina in endophytic and soil-dwelling habitats

Illumina amplicon sequencing of soil in a temperate pine forest in the southeastern United States detected an abundant, nitrogen (N)-responsive fungal genotype of unknown phylogenetic affiliation. Two isolates with ribosomal sequences consistent with that genotype were subsequently obtained. Examination of records in GenBank revealed that a genetically similar fungus had been isolated previously as an endophyte of moss in a pine forest in the southwestern United States. The three isolates were characterized using morphological, genomic, and multilocus molecular data (18S, internal transcribed spacer [ITS], and 28S rRNA sequences). Phylogenetic and maximum likelihood phylogenomic reconstructions revealed that the taxon represents a novel lineage in Mucoromycotina, only preceded by Calcarisporiella, the earliest diverging lineage in the subphylum. Sequences for the novel taxon are frequently detected in environmental sequencing studies, and it is currently part of UNITE’s dynamic list of most wanted fungi. The fungus is dimorphic, grows best at room temperature, and is associated with a wide variety of bacteria. In this paper, a new monotypic genus, Bifiguratus, is proposed, typified by Bifiguratus adelaidae.

54 ENVIRONMENTAL SCIENCES↗