In-situ confocal Raman monitoring of evaporation process during protein crystallization

Vongsvivut, Jitraporn, Watanabe, Shu, Noda, Kohki, Sato, Hidetoshi and Tashiro, Hideo 2008, In-situ confocal Raman monitoring of evaporation process during protein crystallization, Science Asia, vol. 34, no. 1, pp. 400-408, doi: 10.2306/scienceasia1513-1874.2008.34.400.

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Title In-situ confocal Raman monitoring of evaporation process during protein crystallization
Author(s) Vongsvivut, Jitraporn
Watanabe, Shu
Noda, Kohki
Sato, Hidetoshi
Tashiro, Hideo
Journal name Science Asia
Volume number 34
Issue number 1
Start page 400
End page 408
Total pages 9
Publisher Science Society of Thailand under the Patronage of His Majesty the King
Place of publication Bangkok, Thailand
Publication date 2008
ISSN 1513-1874
Summary We have introduced an in-situ Raman monitoring technique to investigate the crystallization process inside protein drops. In addition to a conventional vapour-diffusion process, a novel procedure which actively stimulates the evaporation from a protein drop during crystallization was also evaluated, with lysozyme as a model protein. In contrast to the conventional vapour-diffusion condition, the evaporation-stimulated growth of crystals was initiated in a simple dehydration scheme and completed within a significantly shorter time. To gain an understanding of crystallization behaviours under the conditions with and without such evaporation stimulation, confocal Raman spectroscopy combined with linear regression analysis was used to monitor both lysozyme and HEPES buffer concentrations in real time. The confocal measurements having a high spatial resolution and good linear response revealed areas of local inhomogeneity in protein concentration when the crystallization started. The acquired concentration profiles indicated that (1)ÿthe evaporation-stimulated crystallization proceeded with protein concentrations lower than those under conventional vapour diffusion, and (2)ÿcrystals under the evaporation-stimulated condition were noticeable within an early stage of crystallization before the protein concentration approached its maximum value. The HEPES concentration profiles, on the other hand, increased steadily towards the end of the process regardless of the conditions used for crystallization. In particular, the observed local inhomogeneities specific to protein distribution suggested an accumulation mechanism of protein molecules that initiates the nucleation of crystals.
Notes This article was featured on the cover of Science Asia 34(1)
Language eng
DOI 10.2306/scienceasia1513-1874.2008.34.400
Field of Research 030101 Analytical Spectrometry
Socio Economic Objective 970103 Expanding Knowledge in the Chemical Sciences
HERDC Research category C1.1 Refereed article in a scholarly journal
Copyright notice ©2008, Science Society of Thailand under the Patronage of His Majesty the King
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Document type: Journal Article
Collection: School of Life and Environmental Sciences
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