Proteomics Approach for Predicting Separation Behavior of a Mixture of Proteins During Downstream Purification

Proteomics approach for predicting separation behavior of a mixture of proteins during downstream purification

Ryan Swanson¹, Ruo Xu², Dan Nettleton² and Charles E. Glatz¹

¹Department of Chemical and Biological Engineering, Iowa State University, Ames, IA 50011

²Department of Statistics, Iowa State University, Ames, IA 50011

Recombinant protein production has many applications spanning a wide range of scientific fields, with downstream purification being the most costly part of the overall process. One reason for this stems from a lack of knowledge of the selected host cell protein's (HCP) separation behavior during downstream purification. The process of selecting the downstream purification method(s) as well as the host cell can benefit from an accurate prediction of the HCP separation behavior, thereby reducing the resources needed to investigate both. The HCP separation behavior was characterized using a method where three of the most common protein properties involved in downstream purification were obtained by aqueous two-phase partitioning or ATPS (surface hydrophobicity) and 2-dimensional electrophoresis or 2DE (pI or isoelectric point and molecular weight). Once the proteins had been characterized or “mapped-out” by the three properties, a multivariate model was generated with each characterization property used as a predictor variable and separation behavior as the response variables. The accuracy of predicting a proteins separation behavior using this 3-dimensional characterization method (ATPS + 2DE) had been previously verified for ion exchange chromatography using a set of known model proteins. Here results for hydrophobic interaction chromatography will be added, again using a set of known proteins. This talk will additionally discuss the extension of alternative statistical models for predicting the separation behavior of selected downstream purification methods from 3D characterization of a complex HCP extract (transgenic corn).