Buffer and additive thermofluor screening of wild type human interferon gamma and mutant proteins

•Buffer and additive screening of highly prone to aggregation proteins.•Identification of the best buffer for hIFNγ and hIFNγ mutants.•Identification of the best additives for hIFNγ and hIFNγ mutants.•Identification of the best storage conditions for hIFNγ and hIFNγ mutants.•Step towards further investigation and application of the target protein therapeutics.

Human interferon gamma (hIFNγ) plays a key role in the immune system and therefore this cytokine has many current and future therapeutic applications. hIFNγ is well known with its aggregation propensity and despite its clinical use, there is not much data about the stabilization of hIFNγ preparations. Nowadays, substantial evidence indicates that the pathogenesis of many autoimmune diseases is related to overproduction of hIFNγ. In this regard we have developed inactive hIFNγ analogues to act as receptor antagonists of the endogenous hIFNγ. Since they show even higher tendency for aggregation than the wild type protein, we designed two-step thermofluor screen of 61 buffer conditions to identify the best storage solution for all investigated proteins to be used in the form of research samples or biopharmaceuticals. Tris buffer pH 8.0 supplemented with NaCl and trehalose/betaine/glycerol as additives showed to be the most appropriate choice ensuring high solubility and thermal stability of both hIFNγ and its mutants.