Effects of impurities on membrane-protein crystallization in different systems

Journal Article · Wed Sep 16 00:00:00 EDT 2009 · Acta Crystallographica. Section D: Biological Crystallography

DOI:https://doi.org/10.1107/s0907444909029163· OSTI ID:1625663

Kors, Christopher A. ^[1]; Wallace, Ellen ^[2]; Davies, Douglas R. ^[2]; Li, Liang ^[3]; Laible, Philip D. ^[1]; Nollert, Peter ^[2]

---

1. Argonne National Lab. (ANL), Argonne, IL (United States). Biosciences Division
2. deCODE biostructures, Bainbridge Island, WA (United States)
3. Univ. of Chicago, IL (United States). Inst. for Biophysical Dynamics. Dept. of Chemistry

When starting a protein-crystallization project, scientists are faced with several unknowns. Amongst them are these questions: (i) is the purity of the starting material sufficient? and (ii) which type of crystallization experiment is the most promising to conduct? The difficulty in purifying active membrane-protein samples for crystallization trials and the high costs associated with producing such samples require an extremely pragmatic approach. Additionally, practical guidelines are needed to increase the efficiency of membrane protein crystallization. In order to address these conundrums, the effects of commonly encountered impurities on various membrane-protein crystallization regimes have been investigated and it was found that the lipidic cubic phase (LCP) based crystallization methodology is more robust than crystallization in detergent environments using vapor diffusion or microbatch approaches in its ability to tolerate contamination in the forms of protein, lipid or other general membrane components. LCP-based crystallizations produced crystals of the photosynthetic reaction center (RC) of Rhodobacter sphaeroides from samples with substantial levels of residual impurities. Crystals were obtained with protein contamination levels of up to 50% and the addition of lipid material and membrane fragments to pure samples of RC had little effect on the number or on the quality of crystals obtained in LCP-based crystallization screens. If generally applicable, this tolerance for impurities may avoid the need for samples of ultrahigh purity when undertaking initial crystallization screening trials to determine preliminary crystallization conditions that can be optimized for a given target protein.

• Research Organization: Argonne National Laboratory (ANL), Argonne, IL (United States)
• Sponsoring Organization: USDOE Office of Science (SC), Basic Energy Sciences (BES). Chemical Sciences, Geosciences & Biosciences Division
• Grant/Contract Number: AC02-06CH11357
• OSTI ID: 1625663
• Journal Information: Acta Crystallographica. Section D: Biological Crystallography, Vol. 65, Issue 10; ISSN 0907-4449
• Publisher: International Union of Crystallography
• Country of Publication: United States
• Language: English

Related Subjects

37 INORGANIC, ORGANIC, PHYSICAL, AND ANALYTICAL CHEMISTRY
59 BASIC BIOLOGICAL SCIENCES
Biochemistry & Molecular Biology
Biophysics
Crystallography