An investigation of the effects of varying pH on protein crystallization screening

Chen, Ruiqing; Cheng, Qing-Di; Chen, Jingjie; Sun, Da-Shan; Ao, Liang-Bo; Li, Dawei; Lu, Qi; Yin, Da‐Chuan

doi:10.1039/c6ce02136k

Cited by 15 publications

(11 citation statements)

References 31 publications

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“…Moreover, due to the impurity of the proteins in solution nucleation and crystallisation from fermentation broths are rarely successful. In addition, the crystallisation of protein is also influenced by pH (Chen et al, 2017), iron strength (McPherson, 2004), concentration of PEG (McPherson, 1976), magnetic fields (Hou and Chang, 2008) and etc.…”

Section: Introductionmentioning

confidence: 99%

Continuous protein crystallisation platform and process: Case of lysozyme

Yang

Peczulis

Inguva

et al. 2018

Chemical Engineering Research and Design

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In this work, we designed and built a continuous crystallisation oscillatory flow platform. The lysozyme crystallisation behaviours were investigated at concentrations from 30 to 100 mg/mL, under oscillatory conditions with amplitude (0) from 10 to 25 mm and frequency () from 0.05 to 0.25 Hz in a batch oscillatory flow crystallisation platform. The nucleation rate increased with increase in concentration of initial lysozyme solution, and was also found to increase with increase in shear rate. By learning the thermodynamics and kinetics of lysozyme crystallisation in batch oscillatory flow, the batch crystallisation process was successfully transferred to a continuous oscillatory flow crystallisation process. The equilibrium state of continuous crystallisation reached at residence time 200 min, and the final product crystals shape and size were consistent during the continuous process. This work demonstrates the feasibility of oscillatory flow based platforms for the development of continuous protein crystallisation as for downstream bioseparation.

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Section: Introductionmentioning

confidence: 99%

Continuous protein crystallisation platform and process: Case of lysozyme

Yang

Peczulis

Inguva

et al. 2018

Chemical Engineering Research and Design

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“…Another important factor which strongly affects insulin and protein crystallinity in general is the crystallization pH, as this has been established by several earlier studies [136][137][138][139]. Within a wide pH range, protein molecules may modify in various ways, leading, for example, to partial amino acid neutralization, disrupting the formation of salt bridges between protein molecules, and thus decreasing the crystallization rate.…”

Section: Distinct and Novel Hi Polymorphs Identified Via Xrpdmentioning

confidence: 99%

Applications of X-ray Powder Diffraction in Protein Crystallography and Drug Screening

et al. 2020

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Providing fundamental information on intra/intermolecular interactions and physicochemical properties, the three-dimensional structural characterization of biological macromolecules is of extreme importance towards understanding their mechanism of action. Among other methods, X-ray powder diffraction (XRPD) has proved its applicability and efficiency in numerous studies of different materials. Owing to recent methodological advances, this method is now considered a respectable tool for identifying macromolecular phase transitions, quantitative analysis, and determining structural modifications of samples ranging from small organics to full-length proteins. An overview of the XRPD applications and recent improvements related to the study of challenging macromolecules and peptides toward structure-based drug design is discussed. This review congregates recent studies in the field of drug formulation and delivery processes, as well as in polymorph identification and the effect of ligands and environmental conditions upon crystal characteristics. These studies further manifest the efficiency of protein XRPD for quick and accurate preliminary structural characterization.

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“…Owing to the fact that the solubility of pro-teins is lowest at the isoelectric point, the risk for the crystallization solution entering the labile zone at this point is the highest, leading to precipitates rather than a controlled crystallization. In addition, a study by Chen [17] showed that the pH should be higher from the pI within the stable pH range for acidic proteins such as L-asparaginase II [18]. Therefore, the crystallization solutions were prepared by using higher pH buffers in the following trials.…”

Section: Preliminary Determination Of Crystallization Conditionsmentioning

confidence: 99%

Purification of Recombinant L‐Asparaginase II Using Solvent‐Freeze‐Out Technology

Huang

et al. 2018

Chem Eng & Technol

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The purification of recombinant L‐asparaginase II from an Escherichia coli fermentation broth was carried out by using solvent‐freeze‐out technology on the lab scale. Preliminary crystallization conditions were determined by using batch crystallization screening. The purification of L‐asparaginase II was performed afterwards. Protein crystals with rhombic shape were obtained from the solvent‐freeze‐out crystallization using ethanol. The purity of the crystals was evaluated with sodium dodecyl sulfate polyacrylamide gel electrophoresis (SDS‐PAGE) together with UV spectrophotometry. It was found that DNA could be mostly removed by using solvent‐freeze‐out crystallization. Furthermore, the enzymatic activity was preserved.

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An investigation of the effects of varying pH on protein crystallization screening

Cited by 15 publications

References 31 publications

Continuous protein crystallisation platform and process: Case of lysozyme

Continuous protein crystallisation platform and process: Case of lysozyme

Applications of X-ray Powder Diffraction in Protein Crystallography and Drug Screening

Purification of Recombinant L‐Asparaginase II Using Solvent‐Freeze‐Out Technology

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