... understanding life in molecular detail

Prof Peter Knowles

Copper Oxidases; Free Radicals; Self-Assembly; Membranes

We are sad to announce that Professor Peter Knowles passed away in September 2015.

An obituary of Peter, written by Prof Mike McPherson can be found on the history pages of the web site.

Current major projects include:

Research interests

 

Structure and function of copper-containing oxidases

The research involves close collaboration between several members of academic staff in the Astbury Centre (Knowles, McPherson, Phillips, Parsons and Wilmot) and the School of Chemistry (Halcrow). Our approach is interdisciplinary and embraces Molecular and Cell Biology, Enzymology, Protein Crystallography, Spectroscopy, Mechanistic Studies and Bio-Inorganic Systems. We have a long-established collaboration with Montana State University (Dr David Dooley) which gives access to other advanced spectroscopic facilities.

Self assembly of proteins and peptides in model membranes

The research involves study of the recognition and self organisation behaviour of proteins and peptides in fluid lipid bilayer membranes tethered to surfaces. This interdisciplinary research involves collaboration between molecular and cell biologists, physiologists, biochemists, chemists, physicists and electronic engineer and forms part of the growing Nanotechnology programme at Leeds.

Research projects

l Copper oxidases

Research focuses on two enzyme classes which oxidise primary alcohols and primary amines.

Galactose oxidase catalyses the stereo-specific oxidation of a variety of sugars and other alcohols. We determined the structure of galactose oxidase at 1.7� in 1991 and cloned, sequenced and expressed the gene. This provides us with a sound structural basis for understanding the molecular basis for catalysis by the enzyme using site-directed mutagenesis, x-ray crystallography, stopped flow kinetics and spectroscopy (optical spectroscopy, EPR, resonance Raman). Galactose oxidase is novel in that it utilises a protein-derived tyrosine free radical to effect catalysis. We are studying the biogenesis of this 'in-built' cofactor and other post-translational processing events in the protein. Bio-inorganic modelling is being used to understand the chemistry of these processing events and to produce a functional catalytic model to galactose oxidase.

Amine oxidases catalyse the stereo-specific oxidation of biogenic amines like dopamine and histamine. Amine oxidase protein has recently been shown to be identical with vascular adhesion protein involved as a receptor in lymphocyte trafficing. The connection between the receptor and catalytic functions of the protein are not at present understood.

We determined the first structure of an amine oxidase in 1995 and cloned and expressed the gene for E. coli amine oxidase. We have generated a number of site-directed mutational variant forms of the protein and these are helping us understand the molecular basis for catalysis. Structural studies using X-ray crystallography of substrate and inhibitor complexes of the enzyme and of intermediates in the catalytic cycle provides a sound basis for kinetic and spectroscopic studies using the anaerobic stopped flow kinetic, uv/vis and EPR spectroscopic facilities we have established here.

l Self-assembly in model membranes

Molecular recognition, self-assembly and self-organisation are essential processes in cell biology. In order to understand and exploit such processes, an interdisciplinary research centre dedicated to the study of Self Organising Molecular Systems (SOMS) was established in Leeds in 1993. SOMS has its own laboratories where biochemists, chemists, physicists and other scientists work together to solve challenging problems. There is close interaction with the parent departments and with other Research Centres at Leeds, including the Astbury Centre for Structural Biology. SOMS has an international reputation for its studies of liquid crystalline materials, self assembled peptide polymers and lipid bilayer membranes tethered to surfaces. With respect to the SOMS theme area of tethered membranes, we have designed and synthesised organic molecules which activate gold-coated silica surfaces such that fluid ('biological-like') lipid bilayers can be attached. Proteins and ion-channelling antibiotics (eg gramicidin) have been incorporated into these bilayers. This allows structural study by surface interrogation methods (surface plasmon resonance, atomic force microscopy, Fourier transform infra-red) and study of their transport function by conductance and fluorescence methods. There is immense potential for exploiting such technology in biosensor and other biomedical devices.

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Research interests

Structure and function of copper-containing oxidases

The research involves close collaboration between several members of academic staff in the Astbury Centre (Knowles, McPherson, Phillips, Parsons and Wilmot) and the School of Chemistry (Halcrow). Our approach is interdisciplinary and embraces Molecular and Cell Biology, Enzymology, Protein Crystallography, Spectroscopy, Mechanistic Studies and Bio-Inorganic Systems. We have a long-established collaboration with Montana State University (Dr David Dooley) which gives access to other advanced spectroscopic facilities.

Self assembly of proteins and peptides in model membranes

The research involves study of the recognition and self organisation behaviour of proteins and peptides in fluid lipid bilayer membranes tethered to surfaces. This interdisciplinary research involves collaboration between molecular and cell biologists, physiologists, biochemists, chemists, physicists and electronic engineer and forms part of the growing Nanotechnology programme at Leeds.

Research projects

l Copper oxidases

Research focuses on two enzyme classes which oxidise primary alcohols and primary amines.

Galactose oxidase catalyses the stereo-specific oxidation of a variety of sugars and other alcohols. We determined the structure of galactose oxidase at 1.7� in 1991 and cloned, sequenced and expressed the gene. This provides us with a sound structural basis for understanding the molecular basis for catalysis by the enzyme using site-directed mutagenesis, x-ray crystallography, stopped flow kinetics and spectroscopy (optical spectroscopy, EPR, resonance Raman). Galactose oxidase is novel in that it utilises a protein-derived tyrosine free radical to effect catalysis. We are studying the biogenesis of this 'in-built' cofactor and other post-translational processing events in the protein. Bio-inorganic modelling is being used to understand the chemistry of these processing events and to produce a functional catalytic model to galactose oxidase.

Amine oxidases catalyse the stereo-specific oxidation of biogenic amines like dopamine and histamine. Amine oxidase protein has recently been shown to be identical with vascular adhesion protein involved as a receptor in lymphocyte trafficing. The connection between the receptor and catalytic functions of the protein are not at present understood.

We determined the first structure of an amine oxidase in 1995 and cloned and expressed the gene for E. coli amine oxidase. We have generated a number of site-directed mutational variant forms of the protein and these are helping us understand the molecular basis for catalysis. Structural studies using X-ray crystallography of substrate and inhibitor complexes of the enzyme and of intermediates in the catalytic cycle provides a sound basis for kinetic and spectroscopic studies using the anaerobic stopped flow kinetic, uv/vis and EPR spectroscopic facilities we have established here.

l Self-assembly in model membranes

Molecular recognition, self-assembly and self-organisation are essential processes in cell biology. In order to understand and exploit such processes, an interdisciplinary research centre dedicated to the study of Self Organising Molecular Systems (SOMS) was established in Leeds in 1993. SOMS has its own laboratories where biochemists, chemists, physicists and other scientists work together to solve challenging problems. There is close interaction with the parent departments and with other Research Centres at Leeds, including the Astbury Centre for Structural Biology. SOMS has an international reputation for its studies of liquid crystalline materials, self assembled peptide polymers and lipid bilayer membranes tethered to surfaces. With respect to the SOMS theme area of tethered membranes, we have designed and synthesised organic molecules which activate gold-coated silica surfaces such that fluid ('biological-like') lipid bilayers can be attached. Proteins and ion-channelling antibiotics (eg gramicidin) have been incorporated into these bilayers. This allows structural study by surface interrogation methods (surface plasmon resonance, atomic force microscopy, Fourier transform infra-red) and study of their transport function by conductance and fluorescence methods. There is immense potential for exploiting such technology in biosensor and other biomedical devices.

Detailed research programme                  Close ▲