This Small Business Innovation Research (SBIR) Phase I project aims to address a key challenge facing modern proteomics - identification and quantification of low abundant biologically functional proteins in small size, dilute and/or archived samples using mass spectrometry (MS)-based analysis. A combined use of novel chemical biology and sample preparation strategies is proposed to tackle this challenge. The set of specific reagents to be developed composes specific Activity Based Protein Profiling (ABPP) probes, cleavable high affinity solid support conjugation for quantitative target enrichment, and stable isotopic labels for MS identification and quantification. An enabling sample preparation method employed prior to the MS analysis will also be optimized for the ABPP probe labeling. At the completion of the project, including a Phase II study on broader ABPP labeling applications, commercial kits will be made available for life science laboratories that conduct proteomic studies.
The broader impacts of this research are, 1) it is a major step forward in advancing MS-based proteomic studies and biomarker discoveries based on functional low abundance proteins found in small size biospecimen; and 2) the successful commercialization of this technology and associated products will expand the breadth of possible studies using ABPP based quantitative proteomics. The technological and societal benefits of these products will become apparent, when these products enable new biomarkers discoveries, and the newly identified biomarkers are used in addressing fundamental biological questions, accelerating the development of new clinical diagnostic tests and new therapeutic agents.
Omic Biosystems is a newly established company located in Rockville, MD and founded around its exclusive, worldwide license from the University of Maryland for its proprietary Deuterium isobaric Amine Reactive Tag or DiART technology. DiART is an isotope-labeled, cleavable tag that can label proteins and peptides for quantification using mass spectrometry. Omic Biosystems plans to make the DiART reagents available through sales and services to laboratories engaged in or customers interested in mass spectrometry-based quantitative proteomic studies. In addition, Omic Biosystems is actively pursuing developing novel DiART based applications. Now that individual genome sequences are available for human and many other organisms, new knowledge in basic biology and medicine are expected to come largely from studies in proteomics and other ’omics-based life sciences. The primary goal of proteomics studies is the identification and characterization of large numbers of proteins within the proteome, their functions, spatial and temporal distributions in cells and tissues. This NSF SBIR Phase I project was aimed to establish highly specific, efficient and reproducible methods for proteomic studies based on the development of a novel Activity-Based Protein Profiling (ABPP)-DiART probe design coupled with advancements in sample preparation techniques. Highlights of the completed work include: As a result of this NSF SBIR and other sources of funding, Omic Biosystems was able to move further on the course of commercializing the DiART technology for quantitative proteomic studies. We demonstrated, using a simplified protein model, that DiART reagents can be applied in high quality data generation for proteomic analysis, validating the original DiART design. The proof-of-principle results were published in a recent Analytical Chemistry article (Zhang, et. al, 2010). The original design of the ABPP-DiART probe was synthesized successfully. However, we encountered solubility problems with the newly designed probe. Based on this discovery, alternative approaches were proposed for future studies. In collaborating with a number of academic laboratories, we have begun studies in the analysis of complex biological samples using DiART. We have initiated collaborative studies on 1) a novel sample preparation workflow for the analysis of affinity enriched proteins and peptides, and 2) improvement of peptide coverage in mass spectrometry-based protein and peptide identifications by combining new separation technologies with DiART. In summary, we completed a successful proof-of-principle study of DiART and a partially successful investigation on ABPP-DiART technology. This project led us to initiate new investigations on additional DiART related technologies meeting broad proteomic technological needs. The technological and societal benefits of these products supported in part by the funding of this NSF SBIR will become apparent, when these products and methods are employed in biomarker discovery studies, addressing fundamental biological questions and leading to the development of new clinical diagnostic tests and new therapeutic agents. Zhang, J., Wang, Y., and Li, S., Deuterium isobaric Amine-Reactive Tags for Quantitative Proteomics. Anal Chem, 82:7588-7595 (2010)
