Ronald Kluger

Academic Title: Professor

Phone: 416-978-3582

Office: DB 444

Email:

Research Homepage: http://www.chem.utoronto.ca/staff/RHK/kluger_r.html

The study of organic chemistry at the interface of biochemistry provides a view of life processes in terms of the mechanisms of chemistry. Students in our research group can develop knowledge and skills in a broad variety of chemical and biochemical techniques, including organic synthesis, reaction kinetics, spectroscopy, enzymology, and protein chemistry. The objectives of our research are based on utilizing the principles of physical organic chemistry to provide a means of developing biochemical mechanisms and applications. Projects range from physical studies of the reactions of phosphates and organic reaction mechanisms of biochemical models to the design of enzyme inhibitors and reagents for protein modification.

We design and prepare molecules that will react with specific regions of proteins based on the presence of charge and nucleophiles. These site-directed reagents have led to the development of new materials which stabilize proteins and permit them to be used in the development of new materials and drugs. The acyl phosphate monoesters are electrophiles that are anionic, an unusual combination that permits them to react at highly specific sites. We prepare molecules based on examination of the structure of the target protein and then analyze the outcome in the modified protein. We have extended the work to produce proteins covalently linked by defined connections and have examined the effects of the protein-protein interactions.

We have developed methods to produce the intermediates in biochemical reactions of coenzymes. The covalent intermediates correspond in detail to similar enzymic intermediates, which we examine for comparison. The differences in reactivity at every step reveal the contribution of the protein.

We have found that lanthanide ions promote hydroxyl acylation reactions of acyl phosphate monoesters. We are developing the use of this specific combination to modify carbohydrates and nucleotides with site-specificity.

Selective Alteration of Proteins

Chemical alterations permit systematic studies of proteins that would dissociate - these have many applications.

• Hemoglobin-Superoxide Dismutase – Chemical Linkages That Create a Dual-Function Protein. Amer Alagic, Agnieszka Koprianiuk, and Ronald Kluger , J. Am. Chem. Soc. 2005, 33, 8036-804 .
• Conjoined Hemoglobins. Loss of Cooperativity and Protein-Protein Interactions. Nikolai Gourianov and Ronald Kluger Biochemistry 2005 44 14989 – 14999.
• Efficient generation of dendritic arrays of cross-linked hemoglobin: symmetry and redundancy Dongxin Hu and Ronald Kluger  Org. Biomol. Chem. 2008 6 151 – 156.
• PEG-Conjugation Enhances Nitrite Reductase Activity of Native and Cross-Linked Hemoglobin Francine E. Lui, Pengcheng Dong, and Ronald Kluger Biochemistry 2008 47 10773-10780.
• Functional cross-linked hemoglobin bis-tetramers: Geometry and cooperativity Dongxin Hu and Ronald Kluger Biochemistry 2008 47 12551-12561.

Acyl phosphate monoesters as biomimetic reagents

Acyl phosphate esters occur in nature but their use as reagents is just being discovered. They have very useful properties, especially as electrophiles in water.

• Lanthanum-catalyzed aqueous acylation of monosaccharides by benzoyl methyl phosphate, Ian James Gray, Bernhard Westermann, Rui Ren and Ronald Kluger Can. J. Chem . 2006 84 620-624.
• Chelation-controlled regioselectivity in the lanthanum-promoted monobenzoylation of monosaccharides in water . Ian James Gray and Ronald Kluger Carb. Res . 2007 342 1998-2002
• Biomimetic Aminoacylation of Ribonucleotides and RNA with Aminoacyl Phosphate Esters and Lanthanum Salts Svetlana Tzvetkova and Ronald Kluger J. Am. Chem. Soc . 2007 129 15848– 15854
• pKa-Dependent Formation of Amides in Water from an Acyl Phosphate Monoester and Amines Jolanta Wodzinska and Ronald Kluger J. Org. Chem. 2008 734753-4754.

Covalent intermediates in the reactions of thiamin enzymes

Thiamin promotes reactions in patterns that reflect those of related enzymes. The role of the protein is clear if they are compared side by side. There are remarkable differences. Our results have led us to reconsider the role of thiamin in enzyme-catalyzed decarboxylation.

• Deuterium-Labeling as a Test of Intramolecular Hydride Mechanisms in the Fragmentation of N1'-methyl-2-(1-hydroxybenzyl)thiamin Glenn Ikeda and Ronald Kluger Can. J. Chem., 2005 83, 1277-1280.
• Making thiamin work faster: acid promoted separation of carbon dioxide. Qingyan Hu and Ronald Kluger,  J. Am. Chem. Soc . 2005 127 12242-12243.
• Protein-enhanced decarboxylation of the covalent intermediate in benzoylformate decarboxylase – desolvation or acid catalysis? , Ronald Kluger and Daria Yu Bioorg. Chem. 2006 34 337-344 (2006)
• Accelerating Unimolecular Decarboxylation by Pre-Associated Acid Catalysis in Thiamin-Derived Intermediates:  Implicating Brønsted Acids as Carbanion Traps in Enzymes”. Ronald Kluger, Glenn Ikeda, Qingyan Hu, Pengpeng Cao, and Joel Drewry. J. Am. Chem. Soc . 2006 128 15856-15864.
• Thiamin Diphosphate Catalysis: Enzymic and Nonenzymic Covalent Intermediates. Ronald Kluger and Kai Tittmann Chem. Rev. 2008 108 1797-1833.

• Catalyzing separation of carbon dioxide in thiamin diphosphate promoted decarboxylationRonald Kluger and Steven Rathgeber FEBS J. 2008 275 6089-6100.