| Four dehydrin proteins from cold tolerant conifer species Picea wilsonii Mast, Picea meyeri Rehd, Picea korainesis Nakai, and named PicW1、PicW2、PicM、PicK. Those proteins are cold acclimated proteins regulated by the season and contain 5 repetitions of QKA segment. Sequence alignment analysis showed that diversity among the proteins is four sites and seven amino acid residues. Four dehydrin proteins have been crystallized by using the vapour-diffusion method, and we successfully crystallize in some consisting conditions. We used differential scanning calorimetry (DSC) to measure the thermal stability of the proteins. The results showed as follow:1. The crystal of PicW2 and PicK grew in INDEX1-44, a consisting condition of 0.1M HEPES pH 7.5,25%w/v PEG3350. The crystal of PicM grew in INDEX1-22, a consisting condition of 0.8 M Succinic Acid pH 7.0. The crystal of PicW1 grew in SCREEN2, a consisting condition of 0.1 M Sodium chloride,0.1M BICINE pH 9.0,20% v/v Polyethlene glycol monomethyl ether 550.2. The results showed that four proteins have a good thermal stability. PicM is the most stabilized protein, with a denature temperature at 187.08℃, compared to the temperature of PicW2 at 178.79℃, PicK at 163.09℃, and PicW1 at 139.61℃. For the heat capacity, PicW2 is 24.90 J·K-1·g-1, PicW1 is 16.63J·K-1·g-1, PicM is 14.35 J·K-1·g-1, PicK is 2.39 J·K-1·g-1.3. I-TASSER protein molecular simulation results of each protein accounts for that only one of the five K segments formed a classical helix structure, and the others were random coils. The No.13 amino acid mutation of PicW1 was clear different from other three proteins in structure. The No.160 to 163 mutation of PicM was signally different from PicW2 and PicK in construction. Evaluated with the DSC results, the mutation of No.160 to 163 could significantly affects the thermal stability and heat capacity of the proteins, on the other hand, the No.13 amino acid mutation in less. |
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