High level expression, efficient purification, and bioactivity of recombinant human metallothionein 3 (rhMT3) from methylotrophic yeast Pichia pastoris

Metallothionein 3 (MT3) is an important biochemical mediator regulating many physiological and pathophysiological processes including neuron cell protection, privation of reactive oxygen species-induced DNA damage, and protection against light induced retinal damage. In this study, a human gene encoding for MT3 with c-terminal extension of His6-tag was inserted into vector pPICZaA, and overexpressed in Pichia pastoris strain X-33. The rhMT3 was purified by one step Ni+-NTA affinity chromatography yielding 270 mg/L of over 90% purity. Functional analysis of the purified rhMT3 using inductively coupled plasma mass spectrometry demonstrated that it has biological function, binding with metal ions Cd2+, Cu2+ and Zn2+. In summary, the experimental procedure we have developed facilitates production of large amounts of an active rhMT3 for further research and drug development.