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Dr. Carlos Filipe

Professor

Department of Chemical Engineering

Chair of the Department

Department of Chemical Engineering

Expertise:
Applied molecular biology, recombinant protein expression and purification, engineering mRNA for stability and readability.
Areas of Specialization:
Research Clusters:
Email:
Office:
JHE 374B
Phone:
+1 905.525.9140 x 27278

Overview

Representative Example of a Research Project

Recombinant proteins are produced for a variety of applications, such as for enzyme production, therapeutics, proteomics, structural studies, and many other areas. The need for these proteins therefore ranges from the very small scale (such as in proteomics and in assays) to very large scale industrial processes. We developed a very simple method to purify recombinant proteins by fusing a target protein with an intein and an elastin-like polypeptide that only requires NaCl, dithiothreitol, and a syringe filter to isolate the target protein from Escherichia coli lysate (J. Am. Chem. Soc.; 2005; 127(32); 11228-11229). The tripartite

Figure: Tripartite fusion system for simple recombinant purification

This tripartite fusion system enables rapid isolation of the target protein without the need for affinity chromatography for purification or proteases for cleavage of the target protein from the fusion. The elastin-like polypeptide tag imparts reversible phase transition behavior to the tripartite fusion so that the fusion protein can be selectively aggregated in cell lysate by the addition of NaCl. The aggregates are isolated by microfiltration andresolubilized by reversal of the phase transition in low ionic strength buffer. After resolubilizing the fusion protein, the intein is activated to cleave the target protein from the elastin-intein tag, and the target protein is then isolated from the elastin-intein fusion by an additional phase transition cycle. The following diagram shows this sequence of steps.

This work has been highlighted as a technology concentrate in Chemical and Engineering News and in the magazine The Scientist.

Education

  • B.Sc. Catholic Portuguese University (1993)
  • PhD. Clemson University (1999)

Publications

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