scholarly journals Enabling Biocatalysis by High-Throughput Protein Engineering Using Droplet Microfluidics Coupled to Mass Spectrometry

ACS Omega ◽  
2018 ◽  
Vol 3 (2) ◽  
pp. 1498-1508 ◽  
Author(s):  
Xue W. Diefenbach ◽  
Iman Farasat ◽  
Erik D. Guetschow ◽  
Christopher J. Welch ◽  
Robert T. Kennedy ◽  
...  
Keyword(s):  
Mass Spectrometry ◽  
Protein Engineering ◽  
High Throughput ◽  
Droplet Microfluidics

On-chip integration of droplet microfluidics and nanostructure-initiator mass spectrometry for enzyme screening

Lab on a Chip ◽  
2017 ◽  
Vol 17 (2) ◽  
pp. 323-331 ◽  
Author(s):  
Joshua Heinemann ◽  
Kai Deng ◽  
Steve C. C. Shih ◽  
Jian Gao ◽  
Paul D. Adams ◽  
...  
Keyword(s):  
Mass Spectrometry ◽  
High Throughput ◽  
Droplet Microfluidics ◽  
Enzymatic Assay ◽  
Enzyme Screening ◽  
Highly Sensitive ◽  
On Chip

μNIMS, a highly sensitive and high throughput technique for enzymatic assay that integrates droplet microfluidics with nanostructure-initiator mass spectrometry (NIMS).

scholarly journals Droplet Microfluidics-Enabled High-Throughput Screening for Protein Engineering

Micromachines ◽  
2019 ◽  
Vol 10 (11) ◽  
pp. 734 ◽  
Author(s):  
Lindong Weng ◽  
James E. Spoonamore
Keyword(s):  
Protein Engineering ◽  
Directed Evolution ◽  
High Throughput ◽  
Protein Design ◽  
High Throughput Screening ◽  
Fluid Phase ◽  
Droplet Microfluidics ◽  
New Paradigm ◽  
Wide Range ◽  
Challenges And Opportunities

Protein engineering—the process of developing useful or valuable proteins—has successfully created a wide range of proteins tailored to specific agricultural, industrial, and biomedical applications. Protein engineering may rely on rational techniques informed by structural models, phylogenic information, or computational methods or it may rely upon random techniques such as chemical mutation, DNA shuffling, error prone polymerase chain reaction (PCR), etc. The increasing capabilities of rational protein design coupled to the rapid production of large variant libraries have seriously challenged the capacity of traditional screening and selection techniques. Similarly, random approaches based on directed evolution, which relies on the Darwinian principles of mutation and selection to steer proteins toward desired traits, also requires the screening of very large libraries of mutants to be truly effective. For either rational or random approaches, the highest possible screening throughput facilitates efficient protein engineering strategies. In the last decade, high-throughput screening (HTS) for protein engineering has been leveraging the emerging technologies of droplet microfluidics. Droplet microfluidics, featuring controlled formation and manipulation of nano- to femtoliter droplets of one fluid phase in another, has presented a new paradigm for screening, providing increased throughput, reduced reagent volume, and scalability. We review here the recent droplet microfluidics-based HTS systems developed for protein engineering, particularly directed evolution. The current review can also serve as a tutorial guide for protein engineers and molecular biologists who need a droplet microfluidics-based HTS system for their specific applications but may not have prior knowledge about microfluidics. In the end, several challenges and opportunities are identified to motivate the continued innovation of microfluidics with implications for protein engineering.

High‐Throughput Mass Spectrometry Complements Protein Engineering

2021 ◽  
pp. 57-79
Author(s):  
Tong Si ◽  
Pu Xue ◽  
Kisurb Choe ◽  
Huimin Zhao ◽  
Jonathan V. Sweedler
Keyword(s):  
Mass Spectrometry ◽  
Protein Engineering ◽  
High Throughput

154: Integration of TPSA and High-Throughput Mass Spectrometry Data Improves Prostate Cancer Prediction

2007 ◽  
Vol 177 (4S) ◽  
pp. 52-53
Author(s):  
Stefano Ongarello ◽  
Eberhard Steiner ◽  
Regina Achleitner ◽  
Isabel Feuerstein ◽  
Birgit Stenzel ◽  
...  
Keyword(s):  
Prostate Cancer ◽  
Mass Spectrometry ◽  
High Throughput ◽  
Mass Spectrometry Data ◽  
Cancer Prediction

High-Throughput Affinity Selection Mass Spectrometry Using SAMDI-MS to Identify Small-Molecule Binders of the Human Rhinovirus 3C Protease

2021 ◽  
pp. 247255522110232
Author(s):  
Michael D. Scholle ◽  
Doug McLaughlin ◽  
Zachary A. Gurard-Levin
Keyword(s):  
Mass Spectrometry ◽  
Drug Discovery ◽  
High Throughput ◽  
High Throughput Screening ◽  
Human Rhinovirus ◽  
Binding Potential ◽  
Affinity Selection ◽  
Response Curves ◽  
Desorption Ionization ◽  
Self Assembled

Affinity selection mass spectrometry (ASMS) has emerged as a powerful high-throughput screening tool used in drug discovery to identify novel ligands against therapeutic targets. This report describes the first high-throughput screen using a novel self-assembled monolayer desorption ionization (SAMDI)–ASMS methodology to reveal ligands for the human rhinovirus 3C (HRV3C) protease. The approach combines self-assembled monolayers of alkanethiolates on gold with matrix-assisted laser desorption ionization time-of-flight (MALDI TOF) mass spectrometry (MS), a technique termed SAMDI-ASMS. The primary screen of more than 100,000 compounds in pools of 8 compounds per well was completed in less than 8 h, and informs on the binding potential and selectivity of each compound. Initial hits were confirmed in follow-up SAMDI-ASMS experiments in single-concentration and dose–response curves. The ligands identified by SAMDI-ASMS were further validated using differential scanning fluorimetry (DSF) and in functional protease assays against HRV3C and the related SARS-CoV-2 3CLpro enzyme. SAMDI-ASMS offers key benefits for drug discovery over traditional ASMS approaches, including the high-throughput workflow and readout, minimizing compound misbehavior by using smaller compound pools, and up to a 50-fold reduction in reagent consumption. The flexibility of this novel technology opens avenues for high-throughput ASMS assays of any target, thereby accelerating drug discovery for diverse diseases.

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