Linking B-factor and temperature-induced conformational transition

The crystallographic B-factor, also called temperature factor or Debye-Waller factor, has long been used as a surrogate for local protein flexibility. However, the use of the absolute B-factor as a probe for protein motion requires reproducibility and intervalidation against chemical and physical variables. Here we report the investigation of the thermal dependence of the crystallographic B-factor and its correlation with protein conformational changes. We solved the B-factor reproducibility issue at high resolution (1.5 [A]) over a broad temperature range (100 K to 325 K) by protecting crystals with hydrocarbon grease during data collection. We found that the crystallographic protein conformation varies as a function of temperature. Further, the demonstrated that the thermal dependence of B-factor as a function of temperature were similar for all atoms (C, N-amide and side chains), without local variations, indicating lack of correlation between temperature-dependent conformational change and the B-factor. These data indicate a linear correlation of B-factor with temperature due to global rigid body motion.