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Gerdts C.J.,Emerald Biostructures | Gerdts C.J.,Emerald Biosystems | Stahl G.L.,Emerald Biostructures | Napuli A.,Seattle Structural Genomics Center for Infectious Disease | And 14 more authors.
Journal of Applied Crystallography | Year: 2010

The Microcapillary Protein Crystallization System (MPCS) is a microfluidic, plug-based crystallization technology that generates X-ray diffraction-ready protein crystals in nanolitre volumes. In this study, 28 out of 29 (93%) proteins crystallized by traditional vapor diffusion experiments were successfully crystallized by chemical gradient optimization experiments using the MPCS technology. In total, 90 out of 120 (75%) protein/precipitant combinations leading to initial crystal hits from vapor diffusion experiments were successfully crystallized using MPCS technology. Many of the resulting crystals produced high-quality X-ray diffraction data, and six novel protein structures that were derived from crystals harvested from MPCS CrystalCards are reported. © 2010 International Union of Crystallography Printed in Singapore-all rights reserved.


Christensen J.,Emerald Biostructures | Gerdts C.J.,Emerald Biosystems | Clifton M.C.,Emerald Biosystems | Stewart L.,Emerald Biosystems
Acta Crystallographica Section F: Structural Biology and Crystallization Communications | Year: 2011

The MPCS Plug Maker is a microcapillary-based protein-crystallization system for generating diffraction-ready crystals from nanovolumes of protein. Crystallization screening using the Plug Maker was used as a salvage pathway for proteins that failed to crystallize during the initial observation period using the traditional sitting-drop vapor-diffusion method. Furthermore, the CrystalCards used to store the crystallization experiments set up by the Plug Maker are shown be a viable container for long-term storage of protein crystals without a discernable loss of diffraction quality with time. Use of the Plug Maker with SSGCID proteins is demonstrated to be an effective crystal-salvage and storage method.


Luo J.,Centocor Research and Development Inc. | Wu S.-J.,Centocor Research and Development Inc. | Lacy E.R.,Centocor Research and Development Inc. | Orlovsky Y.,Centocor Research and Development Inc. | And 6 more authors.
Journal of Molecular Biology | Year: 2010

Interleukin (IL)-12 and IL-23 are heterodimeric proinflammatory cytokines that share a common p40 subunit, paired with p35 and p19 subunits, respectively. They represent an attractive class of therapeutic targets for the treatment of psoriasis and other immune-mediated diseases. Ustekinumab is a fully human monoclonal antibody (mAb) that binds specifically to IL-12/IL-23p40 and neutralizes human IL-12 and IL-23 bioactivity. The crystal structure of ustekinumab Fab (antigen binding fragment of mAb), in complex with human IL-12, has been determined by X-ray crystallography at 3.0 Å resolution. Ustekinumab Fab binds the D1 domain of the p40 subunit in a 1:1 ratio in the crystal, consistent with a 2 cytokines:1 mAb stoichiometry, as measured by isothermal titration calorimetry. The structure indicates that ustekinumab binds to the same epitope on p40 in both IL-12 and IL-23 with identical interactions. Mutational analyses confirm that several residues identified in the IL-12/IL-23p40 epitope provide important molecular binding interactions with ustekinumab. The electrostatic complementarity between the mAb antigen binding site and the p40 D1 domain epitope appears to play a key role in antibody/antigen recognition specificity. Interestingly, this structure also reveals significant structural differences in the p35 subunit and p35/p40 interface, compared with the published crystal structure of human IL-12, suggesting unusual and potentially functionally relevant structural flexibility of p35, as well as p40/p35 recognition. Collectively, these data describe unique observations about IL-12p35 and ustekinumab interactions with p40 that account for its dual binding and neutralization of IL-12 and IL-23. © 2010 Elsevier Ltd.


Li L.,University of Chicago | Fu Q.,University of Chicago | Kors C.A.,Argonne National Laboratory | Stewart L.,deCODE Biostructures Inc. | And 3 more authors.
Microfluidics and Nanofluidics | Year: 2010

This article presents a plug-based microfluidic system to dispense nanoliter-volume plugs of lipidic cubic phase (LCP) material and subsequently merge the LCP plugs with aqueous plugs. This system was validated by crystallizing membrane proteins in lipidic mesophases, including LCP. This system allows for accurate dispensing of LCP material in nanoliter volumes, prevents inadvertent phase transitions that may occur due to dehydration by enclosing LCP in plugs, and is compatible with the traditional method of forming LCP material using a membrane protein sample, as shown by the successful crystallization of bacteriorhodopsin from Halobacterium salinarum. Conditions for the formation of LCP plugs were characterized and presented in a phase diagram. This system was also implemented using two different methods of introducing the membrane protein: (1) the traditional method of generating the LCP material using a membrane protein sample and (2) post LCP-formation incorporation (PLI), which involves making LCP material without protein, adding the membrane protein sample externally to the LCP material, and allowing the protein to diffuse into the LCP material or into other lipidic mesophases that may result from phase transitions. Crystals of bacterial photosynthetic reaction centers from Rhodobacter sphaeroides and Blastochloris viridis were obtained using PLI. The plug-based, LCP-assisted microfluidic system, combined with the PLI method for introducing membrane protein into LCP, should be useful for minimizing consumption of samples and broadening the screening of parameter space in membrane protein crystallization. © 2009 Springer-Verlag.


Trademark
Emerald Biosystems | Date: 2012-11-15

Protein laboratory information management system software.


Trademark
Emerald Biosystems | Date: 2012-02-16

Reagents for scientific or research use; proteins in raw material and crystallized form and protein arrays for scientific or medical research use; chemical solutions for scientific or research use in industry and science. Laboratory apparatus and computer systems for use in protein purification and crystallography, namely, for purifying and characterizing proteins in crystal form. Services for custom preparation of proteins and reagent products, namely, crystallization of proteins and custom design and development of chemical and biochemical reagents.


Trademark
Emerald Biosystems | Date: 2011-12-13

Diagnostic reagents for scientific or research use. Services for custom preparation of reagent products.


Trademark
Emerald Biosystems | Date: 2011-12-13

Diagnostic reagents for scientific or research use. Services for custom preparation of reagent products.


Trademark
Emerald Biosystems | Date: 2011-01-25

Laboratory mixing and storage holder for securing microcapillary tubes used to facilitate crystal growth for the conduct of crystallization or other structural determination experiments of biological or chemical materials.


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