Effects of size, buffer composition, number and position of tag on purity and yield of purification of histidine‐tagged proteins by immobilized metal affinity chromatography

Abstract

Immobilized metal affinity chromatography (IMAC) is a popular technique used for large‐scale purification of proteins because of its high binding specificity, capacity and relative ease and cost of operation. Despite recent advances in improving the robustness of the technique, several factors still affect the level of purity and yield of proteins during IMAC. Using (His)6‐tagged proteins with comparably high level of expression, we are investigating the effects of factors such as protein size, number and position of tags, species of bivalent ions, buffer composition and commonly used additives on the purity and yield of purification in IMAC. Initial experimental data suggests that size of protein has significant influence on purity and yield of purification of (His)6‐tagged proteins using Ni2+ and Co2+ IMAC.