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Anna V Hine

Anna V Hine

Aston University, UK

Title: ProxiMAX randomization: precision protein engineering

Biography

Biography: Anna V Hine

Abstract

ProxiMAX randomization is the technology that lies behind Isogenica’s Colibra™ offering. It is a defined saturation mutagenesis process that delivers precision control of both identity and relative ratio of amino acids at specified locations within a protein/antibody library. Thus unwanted amino acids such as cysteine and methionine can be eliminated from libraries because no constraints are imposed by the genetic code. Moreover, the process is non-degenerate, which means that encoding DNA libraries are as small as is physically possible.  ProxiMAX relies on a process of saturation cycling comprising ligation, amplification and digestion for each cycle and is the science behind the commercial Colibra™ technology.  Currently focused on antibody libraries but with achieved diversities of >99% (6 & 11 saturated codons) and the potential to generate libraries of up to 1014 components, we contest that ProxiMAX randomization is a vital tool in engineering any protein library of the highest quality. This presentation will examine the development of the ProxiMAX process and give examples of libraries created to date.

 

 

Reference:

 

1.     Ferreira Amaral M M, Frigotto L and Hine A V (2017) Beyond the natural proteome: nondegenerate saturation mutagenesis - methodologies and advantages. Meth. Enyzmol. 585:111-133.

 

2.     Frigotto L, Smith M E, Brankin C, Sedani A, Cooper S E, Kanwar N, Evans D, Svobodova S, Baar C, Glanville J, Ullman C G and Hine A V (2015) Codon-precise, synthetic, antibody fragment libraries built using automated hexamer codon additions and validated through next generation sequencing. Antibodies 4:88-102.

 

3.    Chimonides G F, Behrendt J M, Chundoo E, Bland C, Hine A V, Devitt A, Nagel D A and Sutherland A J (2014) Cellular uptake of ribonuclease A functionalised core–shell silica microspheres. J Mater Chem B, 2:7307-7315.

 

4.    Nagel D, Behrendt J M, Chimonides G F, Torr E E, Devitt A, Sutherland A J and Hine A V  (2014) Polymeric microspheres as protein transduction reagents. Mol. Cell Proteomics, 13:1543-1551.

 

5.    Ashraf M, Frigotto L, Smith M E, Patel S, Hughes M D, Poole A J, Hebaishi H R M, Ullman C G and Hine A V (2013) ProxiMAX randomisation: a new technology for non-degenerate saturation mutagenesis of contiguous codons. Biochem. Soc. Trans. 41:1189-1194.