A5080692017- Biofuel production and bioconversion
- Microbial Metabolic Engineering and Bioproduction
- Bacterial Genetics and Biotechnology
- RNA and protein synthesis mechanisms
- Microbial Community Ecology and Physiology
- Gut microbiota and health
- Receptor Mechanisms and Signaling
- Lipid Membrane Structure and Behavior
- Probiotics and Fermented Foods
- Enterobacteriaceae and Cronobacter Research
- Anaerobic Digestion and Biogas Production
- Adenosine and Purinergic Signaling
- Genomics and Phylogenetic Studies
- Microbial Natural Products and Biosynthesis
- Neuropeptides and Animal Physiology
- Drug Transport and Resistance Mechanisms
- Methane Hydrates and Related Phenomena
- Fungal and yeast genetics research
- Enzyme Production and Characterization
- DNA and Nucleic Acid Chemistry
- Protein Structure and Dynamics
- Fungal Biology and Applications
- Fluoride Effects and Removal
- CRISPR and Genetic Engineering
- Antibiotic Resistance in Bacteria
University of California, Santa Barbara
2016-2023
West Virginia University
2021
Michigan State University
2021
Novo Nordisk Foundation
2014-2017
Technical University of Denmark
2014-2017
Bioengineering Center
2017
National Center for Genetic Engineering and Biotechnology
2017
John Wiley & Sons (United States)
2017
Hudson Institute
2017
Stockholm University
2006-2014
The mechanism by which multispanning helix-bundle membrane proteins are inserted into their target remains unclear. In both prokaryotic and eukaryotic cells, cotranslationally the lipid bilayer. Positively charged residues flanking transmembrane helices important topological determinants, but it is not known whether they act strictly locally, affecting only nearest helices, or can globally, topology of entire protein. Here we found that an Escherichia coli inner protein with four five could...
Simple and reliable DNA editing by uracil excision (a.k.a. USER cloning) has been described several research groups, but the optimal design of cohesive ends for multigene assembly remains elusive. Here, we use two model constructs based on expression gfp a four-gene pathway that produces β-carotene to optimize junctions protocol. By combining cloning with genomic integration technology, demonstrate up six fragments can be assembled in one-tube reaction direct genome high accuracy, greatly...
How do integral membrane proteins evolve in size and complexity? Using the small multidrug-resistance protein EmrE from Escherichia coli as a model, we experimentally demonstrated that evolution of composed two homologous but oppositely oriented domains can occur number steps: An original dual-topology evolves, through gene-duplication event, to heterodimer formed by monomers. This simple evolutionary pathway explain frequent occurrence with an internal pseudo-two-fold symmetry axis plane membrane.
Recent genomic analyses have revealed that anaerobic gut fungi possess both the largest number and highest diversity of lignocellulolytic enzymes all sequenced fungi, explaining their ability to decompose lignocellulosic substrates, e.g., agricultural waste, into fermentable sugars. Despite potential, development engineering methods for these organisms has been slow due complex life cycle, understudied metabolism, challenging culture requirements.
The metabolism of archaeal methanogens drives methane release into the environment and is critical to understanding global carbon cycling. Methanogenesis operates at a very low reducing potential compared other forms respiration therefore many anaerobic environments. Harnessing or altering methanogen has mitigate warming even be utilized for energy applications.Here, we report draft genome sequences isolated Methanobacterium bryantii, Methanosarcina spelaei, Methanosphaera cuniculi,...
In the yeast Saccharomyces cerevisiae, microbial fuels and chemicals production on lignocellulosic hydrolysates is constrained by poor sugar transport. For biotechnological applications, it desirable to source transporters with novel or enhanced function from nonconventional organisms in complement engineering known transporters. Here, we identified functionally screened genes three strains of early-branching anaerobic fungi (Neocallimastigomycota) that encode recently discovered Sugars Will...
Anaerobic fungi and methanogenic archaea are two classes of microorganisms found in the rumen microbiome that metabolically interact during lignocellulose breakdown. Here, stable synthetic co-cultures anaerobic fungus Caecomyces churrovis methanogen Methanobacterium bryantii (not native to rumen) were formed, demonstrating microbes from different environments can be paired based on metabolic ties. Transcriptional changes induced by co-culture evaluated C. across a variety substrates identify...
Benthic nitrogen fixation has been estimated to contribute 15 Tg N year−1 the marine budget. With benthic being largely overlooked in more recent surveys, a refocus on diazotrophy was considered important. Variations nitrogenase activity (acetylene reduction-gas chromatography) tropical lagoon western Indian Ocean (Zanzibar, Tanzania) were monitored over 3-year period (2003–2005) and related cyanobacterial diazotrophic microbial diversity using polyphasic approach. Different patterns...
Topogenic sequences direct the membrane topology of proteins by being recognized and decoded integral translocases. In this paper, we have compared minimal sequence characteristics helical-hairpin, reverse signal-anchor, stop-transfer in bacterial that use either YidC or SecYEG translocases for insertion. We find a stretch composed 3 leucines 16 alanines is required efficient membrane-anchoring M13 procoat protein inserts helical hairpin mechanism, only 19 has detectable ability. Similar...
Bacterial surface display is an attractive technique for the production of cell-anchored, functional proteins and engineering whole-cell catalysts. Although various outer membrane have been used display, easy versatile high-throughput-compatible assay evaluating developing systems missing. Using a single domain antibody (also called nanobody) with high affinity green fluorescent protein (GFP), we constructed system that allows fast, fluorescence-based detection displayed proteins. The hybrid...
Anaerobic fungi (Neocallimastigomycota) are emerging non-model hosts for biotechnology due to their wealth of biomass-degrading enzymes, yet tools engineer these have not been established. Here, we show that the anaerobic gut most GC depleted genomes among 443 sequenced organisms in fungal kingdom, which has ramifications heterologous expression genes as well CRISPR-based genome engineering approaches. Comparative genomic analyses suggest may contain cellular machinery aid sexual...
Engineered cell factories that convert biomass into value-added compounds are emerging as a timely alternative to petroleum-based industries. Although often overlooked, integral membrane proteins such solute transporters pivotal for engineering efficient microbial chassis. Anaerobic gut fungi, adapted degrade raw plant in the intestines of herbivores, potential source valuable biotechnology, yet very little is known about constituents these non-conventional organisms. Here, we mined...
Membrane-associated Cytochromes P450 (P450s) are one of the most important enzyme families for biosynthesis plant-derived medicinal compounds. However, hydrophobic nature P450s makes their use in robust cell factories a challenge. Here, we explore small library N-terminal expression tag chimeras model plant CYP79A1 different Escherichia coli strains. Using high-throughput screening platform based on C-terminal GFP fusions, identify several highly expressing and robustly performing chimeric...
The quaternary structure of the homodimeric small multidrug resistance protein EmrE has been studied intensely over past decade. Structural models derived from both two- and three-dimensional crystals show as an anti-parallel homodimer. However, resolution structures is rather low their relevance for in vivo situation questioned. Here, we have challenged available structural by a comprehensive Trp scanning all four transmembrane helices EmrE. results are close agreement with degree lipid...
Membrane-embedded transporters are crucial for the stability and performance of microbial production strains.Apart from engineering known derived model systems, it is equally important to identify nonconventional organisms that confer advantageous traits biotechnological applications.Here, we transferred genes encoding fluoride exporter (FEX) proteins three strains early-branching anaerobic fungi (Neocallimastigomycota) Saccharomyces cerevisiae.The heterologous localized plasma membrane...
Abstract Biocontainment systems are needed to neutralize genetically modified organisms (GMOs) that pose ecological threats outside of controlled environments. In contrast, benign selection markers complement GMOs with reduced fitness. Benign agents serve as alternatives antibiotics, which costly and risk spread antibiotic resistance. Here, we present a yeast biocontainment strategy leveraging engineered fluoride sensitivity DNA vectors enabling use agent. The system addresses the scarcity...
G protein-coupled receptors (GPCRs) have long been shown to exist as oligomers with functional properties distinct from those of the monomeric counterparts, but driving factors oligomerization remain relatively unexplored. Herein, we focus on human adenosine A 2A receptor (A R), a model GPCR that forms both in vitro and vivo. Combining experimental computational approaches, discover intrinsically disordered C-terminus R drives homo-oligomerization. The formation declines progressively...