Abstract
Interferon-tau (IFN-tau) is a novel type I IFN that was originally identified as a pregnancy recognition hormone. IFN-tau shares all of the biological properties of other type I IFNs including antiviral activity and antiproliferative activity through induction of the cell cycle inhibitor gene product p21WAF1. It is a promising therapy for cancers, viral infections, and for autoimmune disorders such as multiple sclerosis, without the adverse side effects associated with IFN-alpha and IFN-beta. Here, we describe novel growth and induction conditions for the expression of functionally active and uniformly 15N-labeled IFN-tau from Pichia pastoris in a minimal media for use in initial 2D- and 3D-NMR studies in solution. Purified 15N-IFN-tau was homogenous, as determined by sodium dodecyl sulfate polyacrylamide gel electrophoresis (SDS-PAGE) and MALDI-TOF mass spectrometer (MS), and retained full biological activity. MS analysis confirmed uniform isotopic labeling of IFN-tau with 15N incorporation exceeding 99%. Circular dichroism (CD) as well as 1D-NMR and 15N-1H heteronuclear single quantum coherence (HSQC) spectra confirmed that purified 15N-labeled IFN-tau has a stable secondary structure. Besides providing a route for isotope labeling of IFN-tau, our procedure may be useful for the expression and purification of other proteins that are difficult to obtain in Pichia pastoris grown in minimal media.
Get full access to this article
View all access options for this article.