title: Cleaning protocols for crystallization robots: preventing protease contamination.
abstract: The protease in the commonly used commercial low-foam enzyme cleaner Zymit cannot be completely blocked by EDTA, a widely used inhibitor of metalloproteases, at concentrations of up to 5 mM. Severe protein degradation was observed in crystallization drops after EDTA-containing wash steps unless residual Zymit protease was removed with NaOH at a concentration of at least 0.1 M. Wash steps with 0.1% SDS were also ineffective in completely removing the remaining Zymit activity. Protocols including wash steps with at least 0.1 M NaOH, as for example specified in the original ZENM protocol, are recommended to completely deactivate Zymit protease activity.
authors.affiliation: Division of Biological Chemistry (Biocenter), Medical University of Innsbruck, Innrain 80, 6020 Innsbruck, Austria.
authors.forename: Andreas
authors.initials: A
authors.lastname: Naschberger
authors.affiliation: Division of Biological Chemistry (Biocenter), Medical University of Innsbruck, Innrain 80, 6020 Innsbruck, Austria.
authors.forename: Barbara G
authors.initials: BG
authors.lastname: Fürnrohr
authors.affiliation: Division of Biological Chemistry (Biocenter), Medical University of Innsbruck, Innrain 80, 6020 Innsbruck, Austria.
authors.forename: Theresia
authors.initials: T
authors.lastname: Dunzendorfer-Matt
authors.affiliation: Art Robbins Instruments, 1293 Mountain View Alviso Road, Suite D, Sunnyvale, CA 94089, USA.
authors.forename: Christopher A
authors.initials: CA
authors.lastname: Bonagura
authors.affiliation: Art Robbins Instruments, 1293 Mountain View Alviso Road, Suite D, Sunnyvale, CA 94089, USA.
authors.forename: David
authors.initials: D
authors.lastname: Wright
authors.affiliation: Division of Biological Chemistry (Biocenter), Medical University of Innsbruck, Innrain 80, 6020 Innsbruck, Austria.
authors.forename: Klaus
authors.initials: K
authors.lastname: Scheffzek
authors.affiliation: Department of Genetic Epidemiology, Medical University of Innsbruck, Schöpfstrasse 41, 6020 Innsbruck, Austria.
authors.forename: Bernhard
authors.initials: B
authors.lastname: Rupp
mesh_terms: D057205:Automation, Laboratory; D003460:Crystallization; D018360:Crystallography, X-Ray; D004866:Equipment Contamination; D010447:Peptide Hydrolases / Q000737:chemistry*; D011480:Protease Inhibitors / Q000737:chemistry*; D012371:Robotics
chemical_list: D011480:Protease Inhibitors; D010447:Peptide Hydrolases
keywords: Zymit; cleaning protocol; crystallization robotics; protease; protease inhibitor