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Pfuhl Group

The Pfuhl group studies the structures of proteins and their dynamics and interactions using NMR spectroscopy and other biophysical techniques. We have two main areas of interest: The regulation of muscle contraction and what happens in muscle and cardiovascular disease and the regulation of protein kinases and their partner proteins in cancer. We study the relevant proteins to understand the molecular mechanisms that lead to disease and try to harness this understanding to support the development of new therapies.

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Mark Pfuhl

Mark Pfuhl

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Structure of the human cardiac myosin regulatory light chain in complex with a fragment of the heavy chain (blue).

Structure and function of cardiac myosin binding protein C (cMyBP-C)

In collaboration with the Gautel & Kampourakis groups we are investigating the conformation, dynamics and interactins of several portions of the enigmatic cMyBP-C. This protein interacts with several parts of the myosin heavy and light chains. Some of these interactions are regulated by phosphorylation and we are investigating the conformational changes in the protein induced by phosphorylation.

NMR spectrum of human titin kinase.

Titin kinase

Titin kinase plays an important role in the regulation of protein homeostasis in cardiac muscle. It is speculated that it is activated mechanical force and that this activation allows signalling to the nucleus providing a direct link from the sarcomere to protein synthesis. The activation of titin kinase has so far only been explored by simulations and the aim of the project is to use NMR spectroscopy to explore the activated state of titin kinase experimentally in collaboration with the Gautel group.

Crystal structure of NMRK1

NADH metabolism in the heart

The enzymes NMRK1 and NMRK2 are key players in a stress induced recovery pathway for NADH. While NMRK1 is ubiquitously expressed, NMRK2 is only detected in skeletal and cardiac muscle. In both tissues it is highly overexpressed under stress or after strenuous exercise. In collaboration with the Hughes group we are exploring the function of this protein by a combination of in vitro and in vivo methods.

Interaction of phosphorylated TACC3 with the clathrin heavy chain.

Structure and function of Aurora-A kinase and associated proteins

In collaboration with the Bayliss group (Leeds) we are exploring the regulation and activity of Aurora-A kinase and its associated proteins. Aurora-A kinase is an important cell cyle regulator and is a potential target for anti cancer drugs. Aurora-A kinase lacks important regulatory features present in other kinases of the same family which is compensated by the recruitment of a range of regulatory proteins that are often also targets for phosphorylation by Aurora-A. We have studied the structures of some of the associated proteins and how they are modulated by phosphorylation. More recently we started to focus on the flexible N-terminus of the kinase.

Model of one of the two conformations of the Nek2 leucine zipper shown as backbone cartoon (grey) overlaid with the electrostatic surface.

The dynamics of the Nek2 leucine zipper

The cell cycle regulated Nek2 kinase coordiates the centrosome cycle to the cell cycle to ensure even distribution of DNA on the daughter cells. Nek2 malfunction leads to aneuploidy and subsequently cancer. Nek2 is a dimer and the key portion of the dimerization domain is an unusual, ‘undecided’ leucine zipper (LZ). This LZ exists in two conformations that exchange with a rate of about 20 s-1. In collaboration with the Fry group (University of Leicester) we are characterising these two conformations and are investigating if and how this unusual dynamics influences activity of the kinase.

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Our partners

Research in the group funded by the British Heart Foundation and the BBSRC.

BHF

British Heart Foundation

BBSRC

Biotechnology & Biological Sciences Research Council

Group lead

mark-pfuhl

Dr Mark Pfuhl

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