Microseed matrix-screening (rMMS): introduction, theory, practice and a new technique for membrane protein crystallization in LCP
Stefan. A. Kolek, Patrick. D. Shaw Stewart, Bastian Brauning
Douglas Instruments Ltd, Research, Douglas House, East Gartson Hungerford,
RG17 7HD, United Kingdom
Random Microseed
Matrix-Screening (rMMS), where seed crystals are added automatically to random
crystallization screens, is a significant recent breakthrough in protein
crystallization [1]. During the eight years since the method was
published, theoretical understanding of the method has increased [2 - 4], and
several important practical variations of the basic method have emerged [5, 6]. We will briefly describe some of these
variations, including cross-seeding, and introduce a novel method of making LCP
seed stocks by scaling up LCP crystallization conditions. We will also describe a method of generating
seed gradients across a plate so that the number of crystals in each LCP bolus
can be varied, with a practical example.
[1] D'Arcy, A., Villard,
F, and Marsh, M. "An automated microseed matrix-screening method for
protein crystallization." Acta Crystallographica Section D: Biological
Crystallography 63.4 (2007): 550-554.
[2] Shaw Stewart, P. D., Kolek, S. A., Briggs, R. A., Chayen, N.
E., & Baldock, P. F. (2011). Random microseeding: a theoretical and
practical exploration of seed stability and seeding techniques for successful
protein crystallization.Crystal Growth & Design, 11(8), 3432-3441.
[3] D'Arcy, A., Bergfors, T., Cowan-Jacob, S. W., & Marsh, M. (2014). Microseed matrix screening for optimization in protein crystallization:
what have we learned?. Acta
Crystallographica Section F: Structural Biology Communications, 70(9), 1117-1126.
[4] Shaw Stewart, P. D, &
Mueller-Dieckmann, J. (2014). Automation in biological crystallization.Acta Crystallographica Section F:
Structural Biology Communications, 70(6),
686-696.
[5] Obmolova, G., Malia, T. J., Teplyakov, A., Sweet, R. W., & Gilliland, G. L. (2014).
Protein crystallization with microseed matrix screening: application to
human germline antibody Fabs. Structural Biology and Crystallization Communications,
70(8).
[6] Abuhammad, A., Lowe, E. D., McDonough, M. A., Shaw Stewart, P. D., Kolek, S. A., Sim, E., & Garman, E. F. (2013). Structure of
arylamine N-acetyltransferase from Mycobacterium
tuberculosis determined by cross-seeding with the homologous protein from M. marinum: triumph over adversity. Acta Crystallographica Section D: Biological
Crystallography, 69(8), 1433-1446.