Effect of temperature programmes on protein crystallisation

Abstract

Varying the temperature has been proven to be beneficial for improving the screening efficiency of protein crystallisation, and thus a crystallisation screening strategy based on this phenomenon can be developed. Such a temperature varying strategy can be applied in practical crystallisation screening, however, there are no guidelines for determining what temperature programme should be utilised. It is therefore necessary to investigate how the temperature programme affects the crystallisation process, so as to help people design a suitable temperature programme. For this purpose, we investigated the effect of temperature programmes on the protein crystallisation (lysozyme, proteinase K, and concanavalin A) that are characterised by different solubility behaviours with respect to temperature. Judging from the reproducibility studies of protein crystallisation with different temperature programmes, we recommend using linear temperature programmes for a moderate time period (24 to 48 h) and a large temperature range according to the properties of the proteins.

Selecting Temperature for Protein Crystallization Screens Using the Temperature Dependence of the Second Virial Coefficient

“…We extracted the data for reproducibility tests of three proteins (lysozyme, proteinase K, and concanavalin A) from our previous publication [34] and carried out a new crystallization reproducibility test of α -chymotrypsinogen A(II). Fig.…”

“…In our recent publications, we have already presented some reproducibility studies at two temperatures (277K and 293K) [34] , which can be used in the current study to show the trend of crystallization success rate versus the temperature. We extracted the data for reproducibility tests of three proteins (lysozyme, proteinase K, and concanavalin A) from our previous publication [34] and carried out a new crystallization reproducibility test of α -chymotrypsinogen A(II). Fig.…”

“… Part of the data ( Fig. 6A , B and D) in this figure were extracted from published results [34] (n = 7). Crystallization methods: hanging drop.…”

Influences of copolymers (Copovidone, Eudragit RL PO and Kollicoat MAE 30 DP) on stability and bioactivity of spray-dried and freeze-dried lysozyme

“…This suggest that some excipients can tolerate heat and absorb heat stress, not the protein included with those excipients.A study byDourado et al (42) showed that Eudragit® L-100 which has a very similar chemical structure to the one used in this study (Kollicoat® MAE 30 DP) can form weak bound conjugates with proteins(38). Kollicoat® MAE 30 DP (Fig1b) contains several methyl groups in its molecular structure.…”

Enhancing Protein Crystallization under a Magnetic Field

“…Therefore, many efforts have been made to increase the sample size and also to decrease the time required. Towards this end, many methods have been proposed, including the grid screen and incomplete factorial approaches [14][15][16], controlling the physical parameters such as the temperature [17,18], hydrodynamic field [19], electric field [20][21][22], magnetic field [23][24][25][26][27][28][29][30][31][32][33][34][35][36][37][38], and electromagnetic field [27,39,40] Despite various trials, obtaining a single crystalline outcome remains a challenge for some proteins owing to their low crystallization ability that prevents screening. In general, experimental parameters such as temperature, pH, and pressure, which are thought to be easily accessible, are selected.…”