Dirk Weinrich

Bio: Dirk Weinrich is an academic researcher from Max Planck Society. The author has contributed to research in topics: Biochip & Protein microarray. The author has an hindex of 7, co-authored 9 publications receiving 1037 citations.

Chemical Strategies for Generating Protein Biochips

Abstract: Protein biochips are at the heart of many medical and bioanalytical applications. Increasing interest has been focused on surface activation and subsequent functionalization strategies for immobilizing these biomolecules. Different approaches using covalent and noncovalent chemistry are reviewed; particular emphasis is placed on the chemical specificity of protein attachment and on retention of protein function. Strategies for creating protein patterns (as opposed to protein arrays) are also outlined. An outlook on promising and challenging future directions for protein biochip research and applications is also offered.

Photochemical Surface Patterning by the Thiol-Ene Reaction

Abstract: The immobilization of proteins on solid substrates while controlling the size and dimensions of the generated patterns is increasingly relevant in biotechnology. Site-specific immobilization and thus control over the orientation of proteins is particularly important because, as opposed to nonspecific adsorption, it generates homogeneous surface coverage and accessibility to the active site of the protein. Consequently, different types of bioorthogonal reactions have been developed to attach proteins site-specifically to surfaces and to control protein patterning. Herein, we report the photochemical coupling of olefins to thiols to generate a stable thioether bond for the covalent surface patterning of proteins and small molecules. This reaction has been applied previously in solution for carbohydrate and peptide coupling. The thiol-ene photoreaction proceeds at close to visible wavelengths (l = 365–405 nm) and in buffered aqueous solutions. As a result of its specificity for olefins, this photoreaction can be considered to be bioorthogonal, unlike other photochemical methods used previously for protein immobilization. To adopt the thiol-ene reaction for the immobilization of biomolecules, surfaces functionalized with thiols and biomolecules derivatized with olefins were prepared (Figure 1). Polyamidoamine (PAMAM) dendrimers were attached covalently to silicon oxide surfaces. An aminocaproic acid spacer was attached to the dendrimers to create distance from the surface. Cystamine was coupled to the spacer, and subsequent reduction of the disulfide yielded the desired thiolterminated surfaces. A liquid layer of terminal-olefinfunctionalized molecules dissolved in ethylene glycol was spread onto these wafers, which were then covered immediately with a photomask. Subsequent irradiation of the surfaces through the photomask led to patterning with adducts of covalently attached thioethers. To establish the method, we photochemically attached the biotin derivative 1 to a thiol-functionalized surface as described above (Figure 1). After the removal of unreacted biotin molecules, the surface was incubated with Cy5-labeled streptavidin (SAv) to produce a SAv-patterned surface. Fluorescence images of the resulting surface (Figure 1) demonstrated that lateral gradients and patterns with micrometer-sized features (5–100 mm) over areas of centimeters in width (Figure 1A) were readily accessible. Figure 1B,C and the fluorescence-intensity profile in Figure 1D show that the patterns have a well-defined shape and are homogeneous over large distances. When prolonged sonication (4 h) and stringent washing were carried our after irradiation, SAv patterns with similar fluorescence intensities were observed, whereas control experiments with biotin that lacked the olefin linker showed no distinctive SAv patterns. These results indicate that the covalent attachment of biotin to the surface occurs specifically through the proposed thiol-ene reaction and that the nonspecific adsorption of biotin is insignificant. Figure 1E shows that the amount of material immobilized can be modified by changing the irradiation time. The procedure reproducibly requires a short irradiation time of 60 s to yield sufficient surface coverage for fabricating dense SAv patterns. To obtain homogeneous fluorescence signals of the patterns, the starting concentration of the solution that is drop cast onto the surface is also important. When the solution of 1 was diluted (to 1 mm), the Cy5-fluorescence intensity decreased considerably. Further dilution (below 500 mm) resulted eventually in disrupted SAv patterns. The application of more concentrated solutions of 1 (> 20 mm) resulted in the saturation of the fluorescence intensity of the SAv patterns. This behavior corresponds well with the effects observed upon varying the irradiation time. Longer irradi[*] Dr. D. N sse, Dr. H. Schroeder, Dr. R. Wacker, Prof. Dr. C. M. Niemeyer Faculty of Chemistry Biological-Chemical Microstructuring Technical University of Dortmund Otto-Hahn-Strasse 6, 44227 Dortmund (Germany) Fax: (+49)231-755-7082 E-mail: christof.niemeyer@tu-dortmund.de

Oriented Immobilization of Farnesylated Proteins by the Thiol-Ene Reaction

Abstract: Anchoring the protein: Proteins were immobilized rapidly under mild conditions by thiol-ene photocoupling between S-farnesyl groups attached to a genetically encodable “CAAX-box” tetrapeptide sequence (A is aliphatic) at the C terminus of the protein and surface-exposed thiols (see scheme). This method enables the oriented covalent immobilization of proteins directly from expression lysates without additional purification or derivatization steps.

Protein Biochips: Oriented Surface Immobilization of Proteins†

Abstract: Substantial progress in biochip technologies has established an efficient and reliable platform for advanced biological and biomedical applications. In particular, the use of protein biochips in high-throughput screens provides high content information. We briefly introduce here recent developments in protein biochip preparation with a special focus on our own work on new methods for protein immobilization and protein microarray fabrication, including the application of the Diels-Alder reaction, Staudinger ligation, 'click' sulfonamide formation, and the photochemical thiol-ene reaction. These chemical methods allow for oriented, site-specific protein conjugation on solid surfaces with high sensitivity and specificity under mild, aqueous conditions.

Chemische Verfahren zur Herstellung von Proteinbiochips

Abstract: Proteinbiochips sind integrale Bestandteile einer wachsenden Zahl medizinischer und bioanalytischer Anwendungen, weshalb das Interesse an Methoden zur Proteinimmobilisierung fur die Herstellung solcher Chips in den letzten Jahren stark gestiegen ist. Dieser Aufsatz gibt einen Uberblick uber chemische Verfahren zur kovalenten und nichtkovalenten Anbindung von Proteinen auf Oberflachen, wobei ein spezielles Augenmerk auf chemische Selektivitat und die Erhaltung der Proteinfunktion bei der Immobilisierung gelegt wird. Ferner werden Strategien zur Herstellung strukturierter Proteinoberflachen umrissen. Abschliesend wird ein Ausblick auf mogliche Entwicklungsrichtungen im Bereich der Proteinbiochipforschung und -anwendung gegeben.

Thiol–Ene Click Chemistry

Abstract: Following Sharpless' visionary characterization of several idealized reactions as click reactions, the materials science and synthetic chemistry communities have pursued numerous routes toward the identification and implementation of these click reactions. Herein, we review the radical-mediated thiol-ene reaction as one such click reaction. This reaction has all the desirable features of a click reaction, being highly efficient, simple to execute with no side products and proceeding rapidly to high yield. Further, the thiol-ene reaction is most frequently photoinitiated, particularly for photopolymerizations resulting in highly uniform polymer networks, promoting unique capabilities related to spatial and temporal control of the click reaction. The reaction mechanism and its implementation in various synthetic methodologies, biofunctionalization, surface and polymer modification, and polymerization are all reviewed.

Bioorthogonal Chemistry: Fishing for Selectivity in a Sea of Functionality

Abstract: The study of biomolecules in their native environments is a challenging task because of the vast complexity of cellular systems. Technologies developed in the last few years for the selective modification of biological species in living systems have yielded new insights into cellular processes. Key to these new techniques are bioorthogonal chemical reactions, whose components must react rapidly and selectively with each other under physiological conditions in the presence of the plethora of functionality necessary to sustain life. Herein we describe the bioorthogonal chemical reactions developed to date and how they can be used to study biomolecules.

Thiol-click chemistry: a multifaceted toolbox for small molecule and polymer synthesis

Abstract: The merits of thiol-click chemistry and its potential for making new forays into chemical synthesis and materials applications are described Since thiols react to high yields under benign conditions with a vast range of chemical species, their utility extends to a large number of applications in the chemical, biological, physical, materials and engineering fields This critical review provides insight into emerging venues for application as well as new mechanistic understanding of this exceptional chemistry in its many forms (81 references)

Functionalizing nanoparticles with biological molecules: developing chemistries that facilitate nanotechnology.

Abstract: Chemistries that Facilitate Nanotechnology Kim E. Sapsford,† W. Russ Algar, Lorenzo Berti, Kelly Boeneman Gemmill,‡ Brendan J. Casey,† Eunkeu Oh, Michael H. Stewart, and Igor L. Medintz*,‡ †Division of Biology, Department of Chemistry and Materials Science, Office of Science and Engineering Laboratories, U.S. Food and Drug Administration, Silver Spring, Maryland 20993, United States ‡Center for Bio/Molecular Science and Engineering Code 6900 and Division of Optical Sciences Code 5611, U.S. Naval Research Laboratory, Washington, D.C. 20375, United States College of Science, George Mason University, 4400 University Drive, Fairfax, Virginia 22030, United States Department of Biochemistry and Molecular Medicine, University of California, Davis, School of Medicine, Sacramento, California 95817, United States Sotera Defense Solutions, Crofton, Maryland 21114, United States