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Measuring Protein Interactions by Microchip Self‐Interaction Chromatography
Author(s) -
García Carlos D.,
Hadley DeGail J.,
Wilson W. William,
Henry Charles S.
Publication year - 2003
Publication title -
biotechnology progress
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 0.572
H-Index - 129
eISSN - 1520-6033
pISSN - 8756-7938
DOI - 10.1021/bp025788z
Subject(s) - chromatography , chemistry , protein–protein interaction , virial coefficient , protein crystallization , lysozyme , function (biology) , hydrophilic interaction chromatography , phase (matter) , crystallization , biological system , high performance liquid chromatography , thermodynamics , biochemistry , physics , organic chemistry , evolutionary biology , biology
The self‐interaction of proteins is of paramount importance in aggregation and crystallization phenomena. Solution conditions leading to a change in the state of aggregation of a protein, whether amorphous or crystalline, have mainly been discovered by the use of trial and error screening of large numbers of solutions. Self‐interaction chromatography has the potential to provide a quantitative method for determination of protein self‐interactions amenable to high‐throughput screening. This paper describes the construction and characterization of a microchip separation system for low‐pressure self‐interaction chromatography using lysozyme as a model protein. The retention time was analyzed as a function of mobile‐phase composition, amount of protein injected, flow rate, and stationary‐phase modification. The capacity factors ( k ′) as a function of crystallizing agent concentration are compared with previously published values for the osmotic second virial coefficient ( B 22 ) obtained by static light scattering, showing the ability of the chip to accurately determine protein‐protein interactions. A 500‐fold reduction in protein consumption and the possibility of using conventional instrumentation and automation are some of the advantages over currently used methodologies for evaluating protein‐protein interactions.