Identification of Antibody Targets for Tuberculosis Serology using High-Density Nucleic Acid Programmable Protein Arrays

Song, Lusheng; Wallstrom, Garrick; Yu, Xiaobo; Hopper, Marika; Duine, Jennifer Van; Steel, Jason C.; Park, Jin; Wiktor, Peter; Kahn, Peter; Brunner, Al; Wilson, Douglas P.; Jenny-Avital, Elizabeth R.; Qiu, Ji; LaBaer, Joshua; Magee, D. Mitchell; Achkar, Jacqueline M.

doi:10.1074/mcp.m116.065953

Cited by 46 publications

(47 citation statements)

References 56 publications

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“…Tags fused to the proteins enable the immobilization via antibodies and an easy purification. [93] A screening process for identification of new antibody responses to the Mycobacterium tuberculosis proteome with about 4,000 tested genes was performed with NAPPA generated microarrays and yielded 8 proteins with tuberculosis biomarker value [94]. The DAPA technology allows the production of replicate protein arrays [95].…”

Section: Applications Of Cell-free Protein Synthesismentioning

confidence: 99%

Application of Cell-Free Protein Synthesis for Faster Biocatalyst Development

2019

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Cell-free protein synthesis (CFPS) has become an established tool for rapid protein synthesis in order to accelerate the discovery of new enzymes and the development of proteins with improved characteristics. Over the past years, progress in CFPS system preparation has been made towards simplification, and many applications have been developed with regard to tailor-made solutions for specific purposes. In this review, various preparation methods of CFPS systems are compared and the significance of individual supplements is assessed. The recent applications of CFPS are summarized and the potential for biocatalyst development discussed. One of the central features is the high-throughput synthesis of protein variants, which enables sophisticated approaches for rapid prototyping of enzymes. These applications demonstrate the contribution of CFPS to enhance enzyme functionalities and the complementation to in vivo protein synthesis. However, there are different issues to be addressed, such as the low predictability of CFPS performance and transferability to in vivo protein synthesis. Nevertheless, the usage of CFPS for high-throughput enzyme screening has been proven to be an efficient method to discover novel biocatalysts and improved enzyme variants.

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Section: Applications Of Cell-free Protein Synthesismentioning

confidence: 99%

Application of Cell-Free Protein Synthesis for Faster Biocatalyst Development

2019

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“…All sera samples were collected with written informed consent with the approval of Institutional Review Boards (IRB) at University of Florida (Gainsville, FL), Arizona State University (Tempe, AZ) and Albert Einstein College of Medicine (Bronx, NY). Detailed sample information was provided in our previous work [17,20]. The sera from guinea pig TB models vaccinated with BCG were kindly provided by Dr. David N. McMurray from Texas A&M College of Medicine [32].…”

Section: Sera Samplesmentioning

confidence: 99%

“…Compared to purified protein microarrays, cell-free protein microarrays are more likely to produce naturally-folded mammalian proteins due to the decreased sample manipulation and use of enhanced cell extracts with native chaperone proteins. Moreover, the use of nucleic acids vastly simplifies the production of custom arrays since any protein can be produced as long as the gene-of-interest is synthesized; for example, arrays can be produced that represent a specific proteome or signaling pathway [17][18][19][20].…”

Section: Introductionmentioning

confidence: 99%

Multiplexed Nucleic Acid Programmable Protein Arrays

Song

Petritis

et al. 2017

Theranostics

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Rationale: Cell-free protein microarrays display naturally-folded proteins based on just-in-time in situ synthesis, and have made important contributions to basic and translational research. However, the risk of spot-to-spot cross-talk from protein diffusion during expression has limited the feature density of these arrays.Methods: In this work, we developed the Multiplexed Nucleic Acid Programmable Protein Array (M-NAPPA), which significantly increases the number of displayed proteins by multiplexing as many as five different gene plasmids within a printed spot.Results: Even when proteins of different sizes were displayed within the same feature, they were readily detected using protein-specific antibodies. Protein-protein interactions and serological antibody assays using human viral proteome microarrays demonstrated that comparable hits were detected by M-NAPPA and non-multiplexed NAPPA arrays. An ultra-high density proteome microarray displaying > 16k proteins on a single microscope slide was produced by combining M-NAPPA with a photolithography-based silicon nano-well platform. Finally, four new tuberculosis-related antigens in guinea pigs vaccinated with Bacillus Calmette-Guerin (BCG) were identified with M-NAPPA and validated with ELISA.Conclusion: All data demonstrate that multiplexing features on a protein microarray offer a cost-effective fabrication approach and have the potential to facilitate high throughput translational research.

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“…Previously, it was shown that proteins displayed on NAPPA arrays are folded properly to interact with known partners 10 ; furthermore, their enzymatic activity was first exploited in 2018, when it was shown that protein kinases displayed on the microarray autophosphorylate 11 . To date, NAPPA methodology has been used for many distinct applications, including biomarker discovery 12,13,14,15,16,17 , protein-protein interactions 10,18 , substrate identification 19 , and drug screening…”

Section: Introductionmentioning

confidence: 99%

Kinase Inhibitor Screening In Self-assembled Human Protein Microarrays

Festa

LaBaer

2019

JoVE

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The screening of kinase inhibitors is crucial for better understanding properties of a drug and for the identification of potentially new targets with clinical implications. Several methodologies have been reported to accomplish such screening. However, each has its own limitations (e.g., the screening of only ATP analogues, restriction to using purified kinase domains, significant costs associated with testing more than a few kinases at a time, and lack of flexibility in screening protein kinases with novel mutations). Here, a new protocol that overcomes some of these limitations and can be used for the unbiased screening of kinase inhibitors is presented. A strength of this method is its ability to compare the activity of kinase inhibitors across multiple proteins, either between different kinases or different variants of the same kinase. Self-assembled protein microarrays generated through the expression of protein kinases by a human-based in vitro transcription and translation system (IVTT) are employed. The proteins displayed on the microarray are active, allowing for measurement of the effects of kinase inhibitors. The following procedure describes the protocol steps in detail, from the microarray generation and screening to the data analysis.

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Identification of Antibody Targets for Tuberculosis Serology using High-Density Nucleic Acid Programmable Protein Arrays

Cited by 46 publications

References 56 publications

Application of Cell-Free Protein Synthesis for Faster Biocatalyst Development

Application of Cell-Free Protein Synthesis for Faster Biocatalyst Development

Multiplexed Nucleic Acid Programmable Protein Arrays

Kinase Inhibitor Screening In Self-assembled Human Protein Microarrays

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