Mutations, post-translational modification, and non-covalent binding factors all play a role in fine tuning polypeptide sequence to final function. Because of this phenomena, populations of proteins annotated to the same sequence nevertheless can display multiple functions within tissues and disease states. Likewise, the same or similar function may be presented by multiple sequences, with subcellular control mechanisms regulating functional diversity. As a consequence, strictly abundance based biomarkers may lack the necessary dynamic range and greater specificity provided by functional based biomarkers, to define the phenotype. Thus, Functional Proteomic techniques such as described here, support a top-down proteomic strategy starting with functional annotation of the structurally intact protein, and ending with sequence and structural annotation.
Compound-centric Displacement Proteomics — An advantaged method to survey small molecule-protein interactions, Swapan Roy, Ph.D
Nature Methods Application Notes
New Chemical Proteomic Methods to Access Drug-protein Interactions
Biotech Support Group LLC, reports on a new technical poster which describes a new enrichment product –PROfessor™, for functional and chemical proteomic applications. Entitled “New New Chemical Proteomic Methods to Access Drug-protein Interactions”, it was presented at the US HUPO Conference in Baltimore, MD, March 10-11, 2013. The poster report describes two important features of the product: 1) a new way to compress protein concentrations in complex samples thereby enriching the low abundance content, and 2) a method called compound-centric displacement proteomics (CCDP) to enrich for the complement interactome of a small molecule or drug. The latter method does not require the surface immobilization of the compound which can distort the orientation and lead to misleading results. “By taking a composite mix of 6 mixed mode surface chemistries, we are able to achieve protein compression when protein loads exceed the binding capacity of the product. Furthermore, the same phenomena – weak binding energy of the proteins to the surface which allows us to achieve compression, also allows subproteomes, the interactomes if you will, to be displaced by the conformational changes associated with drug binding” states Swapan Roy, Ph.D, President & Founder of Biotech Support Group. “With this product, which can be applied universally towards any sample type and any small molecule compound, we envision new chemical proteomic strategies for annotating and correlating drugs to tissue specific isoforms, accessing promiscuity, elucidating mechanism of action, and identifying biomarkers for personalized medicine.” The poster can be viewed at here
Bovine brain homogenate (BBH) and rat brain homogenate (RBH)
Oka, Amita R., Matthew P. Kuruc, Ketan M. Gujarathi, and Swapan Roy. "Functional Proteomic Profiling of Phosphodiesterases Using SeraFILE™ Separations Platform." International Journal of Proteomics 2012 (2012).
Authors Amita R. Oka, Matthew P. Kuruc, Ketan M. Gujarathi, Swapan Roy published an article in the journal International Journal of Proteomics titled, Functional Proteomic Profiling of Phosphodiesterases Using SeraFILE Separations Platform, which illustrates how SeraFILE™ is used to efficiently produce unique subproteomes, with resulting enzyme measurements integrated into molecular profiles. The results demonstrate that the proposed methods provide a means to simplify intersample differences, and to localize proteins attributable to sample-specific responses.
Compound-Centric Displacement Proteomics - An Advantaged Method To Survey Small Molecule-Protein Interactions
US HUPO 2014, Poster: 096
Frontiers in Proteomics: Advancing Biology Through Technology and Computation
The citation is: Abstract functional proteomics relies in part, on the functional or structural features of intact, non-denatured proteins. As such, chemical and affinity-based proteomics can be considered a subset of functional proteomics. Regardless of the evolving terminology, the subject of chemical proteomics is to identify, characterize and quantify the binding interactions of small compounds to proteomes. The consideration of those interactions as functional modulators within the cell is paramount to understanding a potential therapeutic compound's mechanism of action. These same tools and methods also can help survey the promiscuous behavior of compounds towards multiple proteins and posit such behavior as deterministic of either toxicity or efficacy. We describe herein, new tools and methods for this purpose, called compound-centric displacement proteomics (CCDP). Employing a new product -
PROfessor™ which can non-covalently bind proteins, a subset of proteins can be displaced upon introduction of soluble small compounds. Coupled to LC-MS, quantitative metrics of these affinity-eluted sub-proteomes help characterize and identify interacting proteins. These new methods gain efficiencies over prior covalent-based substitution methods and can serve applications in drug target deconvolution, on-target/off-target specificity, and personalized medicine.