The role of cluster formation and metastable liquid - Liquid phase separation in protein crystallization
(2012) In Faraday Discussions 159. p.313-325- Abstract
We discuss the phase behavior and in particular crystallization of a model globular protein (beta-lactoglobulin) in solution in the presence of multivalent electrolytes. It has been shown previously that negatively charged globular proteins at neutral pH in the presence of multivalent counterions undergo a "re-entrant condensation (RC)" phase behavior (Zhang et al., Phys. Rev. Lett., 2008, 101, 148101), i.e. a phase-separated regime occurs in between two critical salt concentrations, c* < c**, giving a metastable liquid-liquid phase separation (LLPS). Crystallization from the condensed regime has been observed to follow different mechanisms. Near c*, crystals grow following a classic nucleation and growth mechanism; near c**, the... (More)
We discuss the phase behavior and in particular crystallization of a model globular protein (beta-lactoglobulin) in solution in the presence of multivalent electrolytes. It has been shown previously that negatively charged globular proteins at neutral pH in the presence of multivalent counterions undergo a "re-entrant condensation (RC)" phase behavior (Zhang et al., Phys. Rev. Lett., 2008, 101, 148101), i.e. a phase-separated regime occurs in between two critical salt concentrations, c* < c**, giving a metastable liquid-liquid phase separation (LLPS). Crystallization from the condensed regime has been observed to follow different mechanisms. Near c*, crystals grow following a classic nucleation and growth mechanism; near c**, the crystallization follows a two-step crystallization mechanism, i.e, crystal growth follows a metastable LLPS. In this paper, we focus on the two-step crystal growth near c**. SAXS measurements indicate that proteins form clusters in this regime and the cluster size increases approaching c**. Upon lowering the temperature, in situ SAXS studies indicate that the clusters can directly form both a dense liquid phase and protein crystals. During the crystal growth, the metastable dense liquid phase is dissolved. Based on our observations, we discuss a nucleation mechanism starting from clusters in the dilute phase from a metastable LLPS. These protein clusters behave as the building blocks for nucleation, while the dense phase acts as a reservoir ensuring constant protein concentration in the dilute phase during crystal growth.
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- author
- Zhang, Fajun ; Roosen-Runge, F. LU ; Sauter, Andrea ; Roth, Roland ; Skoda, Maximilian W.A. ; Jacobs, Robert M.J. ; Sztucki, Michael and Schreiber, Frank
- publishing date
- 2012-11-01
- type
- Contribution to journal
- publication status
- published
- in
- Faraday Discussions
- volume
- 159
- pages
- 13 pages
- publisher
- Royal Society of Chemistry
- external identifiers
-
- scopus:84867952155
- ISSN
- 1359-6640
- DOI
- 10.1039/c2fd20021j
- language
- English
- LU publication?
- no
- id
- 619a2d1c-5c2d-41c1-816b-d16123917e39
- date added to LUP
- 2018-12-17 09:48:36
- date last changed
- 2022-04-18 01:09:01
@article{619a2d1c-5c2d-41c1-816b-d16123917e39, abstract = {{<p>We discuss the phase behavior and in particular crystallization of a model globular protein (beta-lactoglobulin) in solution in the presence of multivalent electrolytes. It has been shown previously that negatively charged globular proteins at neutral pH in the presence of multivalent counterions undergo a "re-entrant condensation (RC)" phase behavior (Zhang et al., Phys. Rev. Lett., 2008, 101, 148101), i.e. a phase-separated regime occurs in between two critical salt concentrations, c* < c**, giving a metastable liquid-liquid phase separation (LLPS). Crystallization from the condensed regime has been observed to follow different mechanisms. Near c*, crystals grow following a classic nucleation and growth mechanism; near c**, the crystallization follows a two-step crystallization mechanism, i.e, crystal growth follows a metastable LLPS. In this paper, we focus on the two-step crystal growth near c**. SAXS measurements indicate that proteins form clusters in this regime and the cluster size increases approaching c**. Upon lowering the temperature, in situ SAXS studies indicate that the clusters can directly form both a dense liquid phase and protein crystals. During the crystal growth, the metastable dense liquid phase is dissolved. Based on our observations, we discuss a nucleation mechanism starting from clusters in the dilute phase from a metastable LLPS. These protein clusters behave as the building blocks for nucleation, while the dense phase acts as a reservoir ensuring constant protein concentration in the dilute phase during crystal growth.</p>}}, author = {{Zhang, Fajun and Roosen-Runge, F. and Sauter, Andrea and Roth, Roland and Skoda, Maximilian W.A. and Jacobs, Robert M.J. and Sztucki, Michael and Schreiber, Frank}}, issn = {{1359-6640}}, language = {{eng}}, month = {{11}}, pages = {{313--325}}, publisher = {{Royal Society of Chemistry}}, series = {{Faraday Discussions}}, title = {{The role of cluster formation and metastable liquid - Liquid phase separation in protein crystallization}}, url = {{http://dx.doi.org/10.1039/c2fd20021j}}, doi = {{10.1039/c2fd20021j}}, volume = {{159}}, year = {{2012}}, }