A5068117164- Microbial Metabolic Engineering and Bioproduction
- RNA and protein synthesis mechanisms
- Viral Infectious Diseases and Gene Expression in Insects
- CRISPR and Genetic Engineering
- Evolution and Genetic Dynamics
- Microbial Natural Products and Biosynthesis
- Carbohydrate Chemistry and Synthesis
- CAR-T cell therapy research
- Bacterial Genetics and Biotechnology
- Bioinformatics and Genomic Networks
- Advanced biosensing and bioanalysis techniques
- Genomics and Phylogenetic Studies
- interferon and immune responses
- Protein Structure and Dynamics
- Glycosylation and Glycoproteins Research
- Innovative Microfluidic and Catalytic Techniques Innovation
- Lymphoma Diagnosis and Treatment
- Algal biology and biofuel production
- Escherichia coli research studies
- Nanowire Synthesis and Applications
- Enzyme Catalysis and Immobilization
- Enzyme Structure and Function
- 14-3-3 protein interactions
- Transgenic Plants and Applications
- Immune cells in cancer
Michigan State University
2014-2019
University of Minnesota
2018-2019
Michigan United
2014-2015
Significance Enzymes find utility as therapeutics and for the production of specialty chemicals. Changing amino acid sequence an enzyme can increase solubility, but many such mutations disrupt catalytic activity. To evaluate this trade-off, we developed experimental system to relative solubility nearly all possible single point mutants two model enzymes. We that tendency a given solubility-enhancing mutation activity depends, among other factors, on how far position is from active site...
In exponentially growing bacteria, expression of heterologous protein impedes cellular growth rates. Quantitative understanding the relationship between and rate will advance our ability to forward engineer important for metabolic engineering synthetic biology applications. Recently, a work described scaling model based on optimal allocation ribosomes translation. This quantitatively predicts linear microbial with no free parameters. With aim validating this model, we have rigorously...
Comprehensive sequence-function mapping involves detailing the fitness contribution of every possible single mutation to a gene by comparing abundance each library variant before and after selection for phenotype interest. Deep sequencing DNA allows frequency reconstruction tens thousands variants in experiment, yet short read lengths current sequencers makes it challenging probe genes encoding full-length proteins. Here we extend scope maps entire protein sequences with modular, universal...
Synthetic metabolic pathways often suffer from low specific productivity, and new methods that quickly assess pathway functionality for many thousands of variants are urgently needed. Here we present an approach enables the rapid parallel determination sequence effects on flux complete gene-encoding sequences. We show this method can be used to determine over 8000 single point mutants a pyrolysis oil catabolic implanted in Escherichia coli. Experimental sequence-function data sets predicted...
The B-cell surface protein CD19 is present throughout the cell life cycle and uniformly expressed in leukemias, making it a target for chimeric antigen receptor engineered immune therapy. Identifying sequence dependence of binding to antibodies empowers fundamental study more tailored development CD19-targeted therapeutics. To identify antibody-binding epitopes on CD19, we screened comprehensive single-site saturation mutation library human extracellular domain mutations detrimental...
CD19-targeted chimeric antigen receptor (CAR) T-cells (CAR19s) show remarkable efficacy in the treatment of relapsed/refractory acute lymphocytic leukemia and Non-Hodgkin's lymphoma. However, use CAR T-cell therapy against CD19-negative hematological cancers solid tumors has been challenging. We propose CD19-fusion proteins (CD19-FPs) to leverage benefits CAR19s while retargeting this validated cellular alternative tumor antigens. demonstrate ability a fusion CD19 extracellular domain (ECD)...
Enzymes are the ultimate entities responsible for chemical transformations in natural and engineered biosynthetic pathways. However, many enzymes suffer from suboptimal functional expression due to poor intrinsic protein stability. Further, stability enhancing mutations often come at cost of impaired function. Here we demonstrate an automated engineering strategy stabilizing while retaining catalytic function using deep mutational scanning coupled multiple-filter based screening...
Abstract Motivation Deep mutational scanning experiments have enabled the measurement of sequence-function relationship for thousands mutations in a single experiment. The Protein Analysis and Classifier Toolkit (PACT) is Python software package that marries fitness metric given mutation within these to sequence structural features enabling downstream analyses. PACT enables easy development user sharable protocols custom deep as all code modular reusable between protocols. Protocols...
Adenylate cyclase toxin (ACT) is an important Bordetella pertussis virulence factor that not included in current acellular vaccines. We previously demonstrated immunization with the repeat-in-toxin (RTX) domain of ACT elicits neutralizing antibodies mice and discovered first two to neutralize activities by occluding receptor-binding site. Here, we fully characterize these their epitopes. Both bind low nanomolar affinity cross-react homologues produced B. parapertussis bronchiseptica....
A central challenge in the field of metabolic engineering is efficient identification a pathway genotype that maximizes specific productivity over robust range process conditions. Here we review current methods for optimizing pathways living cells. New tools library generation, computational analysis sequence-flux space, and high-throughput screening selection techniques are discussed.
Abstract Deep mutational scanning is now used in directed evolution experiments to quantify the change frequency of a cellular variant mixed population. A key concern extent which enrichment population corresponds fitness metric like relative growth rate or survival percentage. We present here analytical equations converting metrics for plate-based selections. Using isogenic and cultures we show that rates percentages correlate antibiotic These results are important proper interpretation...