Advantages and Disadvantages of His-Tagged Beta-Galactosidase

Abstract

β-Galactosidase is one of the most important biotechnological enzyme used in the dairy industry, pharmacology and in molecular biology. In our laboratory we have overexpressed a recombinant β-galactosidase in Escherichia coli (E. coli). This enzyme differs from its native version (β-GalWT) in that 6 histidine residues have been added to the carboxyl terminus in the primary sequence (β-GalHis), which allows its purification by immobilized metal affinity chromatography (IMAC). In this work we compared the functionality and structure of both proteins and evaluated their catalytic behavior on the kinetics of lactose hydrolysis. We observed a significant reduction in the enzymatic activity of β-GalHis with respect to β-GalWT. Although, both enzymes showed a similar catalytic profile as a function of temperature, β-GalHis presented a higher resistance to the thermal inactivation and evidenced greater half-life time compared to β-GalWT. At room temperature, β-GalHis showed a fluorescence spectrum compatible with a partially unstructured protein however, it exhibited a lower tendency to the thermal-induced unfolding with respect to β-GalWT. Analytical ultracentrifugation experiments demonstrated that the population of β-GalHis molecules exhibited a higher proportion of monomers and a lower proportion of tetrameric species with respect to the His-tag free protein. The impairment of tetramerization may would explain the negative effect of the presence of His-tag on the enzymatic activity. In addition, the present results, analyzed in the context of the available literature, suggest that the effect of the His-tag is protein-specific.

A brief review on basic fundamentals of nanoparticle (NPs)

“…NPs physical property influence its morphological features. Other properties like optical properties include color, light penetration, absorption, reflection capabilities, UV absorption in a solution [6,7] . When NPs are coated into a surface it includes the mechanical properties like elastic, ductile, tensile strength and flexibility.…”

“…What are the advantages of NPs that makes it superior from other drug delivery systems and thus challenges its potential, always hits the reader's/researcher's mind, e.g., [6] 1) enters into tissue or lymphatic systems, biological barriers at molecular level. Diagnosis of cancers, Alzheimer's disease like applications.…”