Ronald Kluger

Professor - Organic and Biological Chemistry

Department of Chemistry, University of Toronto, 80 St. George Street Toronto, Ontario, Canada M5S 3H6

E-mail:

 

Phone: (416)-978-3582

Office: Davenport Building 444

Laboratories: Davenport Building 450 and 451

 

 

Research Interests

o                           The site-specific modification of proteins and enzymes by designed organic compounds is an area of growing importance. We have developed new methods to modify the oxygen-carrier protein, hemoglobin, so it can be used for a number of applications. For example, we have produced multifunctional reagents that cross-link hemoglobin at specific sites, introducing properties that make the material suitable to be used as an alternative for red cells in blood transfusions. We extended the method to include ways to combine cross-linking within a protein and connecting with a second (or additional) protein. This permits the interactions of assembled proteins to be studied in a defined system. We recently have improved the process by applying the CuAAC click procedure that we adapted specifically for coupling of two cross-linked hemoglobin-azides with bis-alkynes

Connecting and cross-linking two hemoglobins

 

lower route is slow - accelerated by enzyme

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  • Decarboxylation mechanisms.

Reversibility. The replacement of a carboxyl group by a proton appears to be a simple matter of forming forming CO2 by breaking a C-C bond. However, catalytic and isotope effect patterns remind us that the CO2 is a powerful electrophile, making the process easily reversible prior to separation of the CO2 molecule from the residual carbanion. We are examining the modes by which this process can be accelerated despite the reversibility. We find that enzymes may function by enhancing the "forward committment", which can be diagnosed by an increased 12C/13C kinetic isotope effect. We also have found that in some cases decarboxylation is accelerated in acidic solutions. We are examining related reactions involving proton transfers and the resulting steric effects that are associated with reorganization.

Bronsted acid prevents reversion

Hydrolytic Route. We have also found that there is a distinct alternative to the loss of CO2: acid-catalyzed addition of water to the carboxyl group and formation of carbonic acid.  Protonated CO2 is an impossible intermediate yet in some cases reactions proceed in highly acidic solutions. This route is analogous to ester hydrolysis and appears to be accessible in systems that provide adequate leaving groups. We are examining the extent to which this process occurs in the many reactions that are reported to undergo acid catalyzed decarboxylation.

hydrolyric decarboxylation

 

 

Reactions are selectively catalyzed by lanthanides (recognition of diol in water)

Aminoacylation of RNA

 

 Learn more about this with a slide presentation: click here

The aminoacyl phosphates are also efficient reagents for producing peptide bonds in water by reaction with amino acid esters. This mimics the non-ribosomal formation of peptides. We are expanding this as a method to produce peptides under conditions that are amenable to biological catalysts.

Selected Publications (by Area)

Overview

The article summarizes the origins of current research as well as earlier activities.

CIC Medal Award Lecture: Molecular keystones: Lessons from bioorganic reaction mechanisms  Ronald Kluger Can. J. Chem. 2006 84 1093-1105

Selective Alteration of Proteins

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

Acyl phosphate monoesters as biomimetic reagents

Acyl phosphate esters occur in nature but their use as reagents has been developed in our lab. They have very useful properties, especially as electrophiles in water.

Catalyzed decarboxylation: thiamin intermediates, addition of water, and reversibility

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 as well as the general process with other catalysts. We have found previously unknown routes for decarboxylation that are also amenable to consideration of how CO2 could be utilized for synthesis.

Group members and their research areas:

Name

Area

Chung-Woo Fung (Technical assistant)

instrumentation, HPLC, mass spectrometry

Yuyang Li Aminoacylation of RNA
Victor Crivianu-Gaita
Dinucleotide aminaocylation
Michael Bielecki Thiamin enzyme intermediates

Adelle Vandersteen

Hydrolytic decarboxylation.

Dr. Scott Mundle 13-C kinetic isotope effects (in Professor Barbara Sherwood Lollar's group) - applications to contaminant hydrology
Graeme Howe Decarboxylation catalysis
Liliana Guevara Opinska Thiamin enzyme mechanisms
Erika Siren Protein-protein conjugation
Serena Singh Efficient formation of bis-tetramers
Aizhou Wang Subunit reaction specificity

GROUP SAFETY NOTICE

 

Last updated October 18, 2013

Presentations hemox manual


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