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. 2007 Aug 31;63(Pt 9):798–801. doi: 10.1107/S1744309107040134

Figure 2.

Figure 2

Crystals of P. furiosus sSTT3 protein prepared from the heat-treated cell lysate and SDS–PAGE analysis of the crystals. (a) Crystals obtained from the peak-top fractions (Fig. 1 ▶, sample 2) after two weeks at 303 K; (b) crystals obtained from the tailing-edge fractions (Fig. 1 ▶, sample 3) after 2 d at 293 K. The scale bars represent 0.1 mm. Crystals were transferred to the corresponding mother liquors for washing, mixed with 5× SDS sample buffer, heated for 5 min at 368 K and then subjected to SDS–PAGE (10–20% gradient gel, Daiichi). The gels were stained with SYPRO Orange protein gel stain (Invitrogen) and were photographed with an LAS-3000 multicolour image analyzer (Fuji Film) using a Blue LED (460 nm) illuminator and a Y515-Di filter. Lane M, prestained markers (Precision Plus protein standards, dual colour, Bio-Rad); lane S, protein solution used for crystallization; lane C, washed crystals. The amount of the markers used was extremely small, so that only the two bands prestained with a fluorescent dye are visible. The 12 kDa band was excised from the CBB-stained gel and subjected to in-gel digestion with trypsin and the eluted peptides were loaded onto LC-ESI-MS/MS (LC Q DECA, Finnigan). The protein was identified by a peptide-fingerprint search using the Mascot software (Matrix Science).