Additional Supporting Information may be found in the online version of this article.

pro2282-sup-0001-suppinfo.tif6153KSupplementary Figure 1. Effects of the Fc-III tag on GFP fluorescence and purification of GFP expressed in eukaryotic cell. (A) The transfected 293T cells with the Fc-III tag; (B) The transfected 293T cells with tag-free GFP; (C) Transfected 293T cells with the Fc-III tagged GFP on its N-terminus; (D) Transfected 293T cells with the Fc-III tagged GFP on its C-terminus. All the fluorescence images were taken under the Nikon fluorescence microscope with the excitation of blue lights, the amplification ratio is 200. (E) The image of the SDS-PAGE of purification the Fc-III tagged GFP from 293T cells. The left Lane, clear supernatant of transfected 293T cells that contains the Fc-III tagged GFP; the middle Lane, molecular maker; the right Lane, the eluates of the Fc-III tagged GFP. (F) Fluorescence intensity of purified WT GFP, Fc-III tagged GFP on its N-terminus and Fc-III tagged GFP on its C-terminus. All the concentrations of proteins are 0.2 mg/ml, excited at 488 nm and examined at 507 nm. All samples were replicated in triplicate, and the data were normalized by the intensity of WT GFP.
pro2282-sup-0002-suppinfo.tif1852KSupplementary Figure 2.The selected MS spectra demonstrating formation of the disulfide bond between Cys74 and Cys146 in oxidized MMCK.(A) The quadruply charged ion formed by the disulfide linkage between tryptic peptides ETPSGFTVDDVIQTGVDNPGHPFIMTVGCVAGDEESYEVFK and GYTLPPHCSR. After reduction, the disulfide-linked peptide generates two fragments: (B) the triply charged ion with mass matching to the tryptic peptide ETPSGFTVDDVIQTGVDNPGHPFIMTVGCVAGDEESYEVFK that contains Cys74, and (C) the doubly charged ion of tryptic peptide GYTLPPHCSR that contains Cys146.
pro2282-sup-0003-suppinfo.tif1077KSupplementary Figure 3. The proposed folding pathway for the tag-free, the His-tagged and the Fc-IIItagged CK based on previous CK folding models [15-16, 20, 28, 32] and the results in this research. The shape size of the reduced or the oxidized CK indicates their relative quantities.

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