Wilmanns Group
Structure and function of protein complexes in biological systems
Figure 1: AGT- (Pex5p)2-AGT complex. Color codes: AGT, yellow, orange; Pex5p, cyan, magenta.
Figure 2: DAPK-CaM complex. Color codes: DAPK, green; CaM, red.
The architecture of the protein interactome in sarcomeric muscle cells. The study of the structure and function of muscle cells is of high scientific interest since many proteins found in these cells, when dysfunctional, are associated with cardiovascular diseases. The overall architecture of ‘sarcomeric units’ inmuscle cells is established by several large protein filament systems such as actin, myosin, nebulin, titin, myomesin and obscurin. We investigate how these proteins are connected and interact with each other, frequentlymediated via small scaffold proteins. Recently, we have determined the structure and function of some key complexes, including telethonin-mediated assembly of the N-terminus of titin (Zou et al., 2006) and C-terminal self-assembly of myomesin (Pinotsis et al., 2009, EMBO J.). Based on these findings, we have been able to unravel molecular key rules for complex formation of sarcomeric proteins with β-sheet domains (Pinotsis et al., 2009, TIBS). Our future focus will be on novel protein interactions within the sarcomeric Z-disk andM-line region, and we will also increasingly investigate novel signalling functions of the protein partners involved.
Activity regulation in protein kinases. The human kinome – the part of the genome that expresses protein kinases – comprises about 600 genes. About 70 protein kinases that phosphorylate either serines or threonines share a common C-terminal autoregulatory domain that is thought to bind calcium/calmodulin (CaM). To investigate the mechanism of activity regulation in these kinases, we first determined the structure of the kinase domain from the giant filament protein titin, in the inhibited apo-conformation (Mayans et al., 1998). Very recently, we were able to determine the structure of another kinase with apoptotic functions – Death Associated Protein Kinase – this time in the presence of CaM (de Diego et al., 2010; figure 2). The structure provides first insight how CaM binding leads to kinase activation by withdrawing the autoregulatory domain from the kinase active site. Our future goal is to complement the ongoing structural studies by in vitro and in vivo functional studies, to decipher underlying, generalmolecularmechanisms that regulate the activity of members of the CaM-dependent protein kinase family. Our ultimate aim is to use these data to promote drug discovery against those kinases which frequently play critical roles in cancer formation and progression.
The architecture of the translocon of peroxisomes. Peroxisomes are cell organelles that allow sequestered metabolic processes that would interfere with other processes that generally take place in the cytosol. Those proteins that are involved in these processes are generally translocated as active and folded targets. We have been able, the first time, to unravel themechanismof the recognition of peroxisome protein targets by the peroxisome import receptor Pex5p, by determining the structure of the cargo binding domain of the receptor in the absence and presence of the cargo protein sterol carrier protein 2 (Stanley et al., 2006) and alanine-glyoxylate aminotransferase (Fodor et al., unpublished; see figure 1). Our present focus is on structural/functional studies of several other protein components of the peroxisomal translocation machinery, including the Pex19p receptor, which recognises proteins that are integrated into the peroxisomalmembrane. Our ultimate goal is putting together data fromknown peroxisomal sub-complexes, to provide insight into the presently unknown overall architecture of the peroxisomal translocon by combined application of a variety of structural biology approaches.
Structural systems biology in M. tuberculosis. During the last three years we have determined the X-ray structures of about ten protein targets, some of them with an already known function and others of unknown function. For instance, we were able to identity Rv2217 as a novel cysteine/lysine dyad acyltransferase, which allows activation of several important protein complexes by lipoylation (Ma et al., 2006).We are planning tomake use of available structural data to investigate functional processes in living mycobacteria by systems biology-oriented approaches (including proteomics, metabolomics, modelling). The ultimate goal of our studies is to make the data available to promote the development of new drugs, vaccines and diagnostic markers against this pathogen.