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Biologische Systeme und Medizin Vortrag: Fr., 10:20–10:40 F-V57
Structure of the Protein Kinase MARK/Par-1: Catalytic and UBA Domain
Saravanan Panneerselvam 1 , Alexander Marx 1 , Eva-Maria Mandelkow 1 ,
Eckhard Mandelkow 1
1 Max-Planck Unit for Structural Molecular Biology, Notkestrasse 85, 22607 Hamburg
MAP/Microtubule affinity regulating kinases (MARKs) are Ser/Thr kinases that phosphorylate
the microtubule-associated proteins tau, MAP2 and MAP4 [1]. Phosphorylation
of the neuronal MAP tau at serine S262 leads to detachment of tau from the surface
of microtubules and to destabilization of microtubules [2]. Phosphorylated tau can aggregate
and form filamentous structures. Formation of paired helical filaments (PHFs)
and neurofibrillary tangles is a hallmark of Alzheimer disease. The closely related kinase
Par-1 is important for the development of cell polarity [3].
The human kinome contains four MARK isoforms. A stable fragment of MARK2,
containing the catalytic and the ubiquitin-associated (UBA) domain, was identified by
limited proteolysis. Structures of the wild type construct and of two mutants, K82R and
T208A/S212A, were determined by x-ray crystallography using synchrotron radiation
of DORIS at DESY, Hamburg [4]. Mutation of K82 to arginine inhibits the kinase
activity by interference with nucleotide binding. T208 is the primary phosphorylation
site in the activation loop which controls access of the substrate. S212 was also found to
be phosphorylated in MARK2 from brain. These two residues were mutated to alanine
to mimic the unphosphorylated state.
All three constructs crystallized in space group P61, in two distinct, but similar crystal
forms, which differ by the length of the c-axis: 106.0 ± 0.37 ˚A (sd, n = 15) for the
double mutant, 99.7 ± 0.23 ˚A (n = 8) for the K82R mutant (both forms observed for
the wild type). The catalytic domain exhibits the typical bi-lobed structure with the
cleft wide opened. Two molecules form a symmetric dimer, with the catalytic clefts
facing each other in the center of the dimer. The UBA domain is attached via a taut
linker to the large lobe of the kinase domain and leans against a hydrophobic patch on
the distal side of the small lobe. The UBA structure is unusual in that the orientation
of its third helix is inverted, relative to previous structures.
Data were collected at the X13 Consortium beamline at HASYLAB (DESY, Hamburg).
We thank W.Rypniewski, M.Perbandt, and J. Müller for help with the beamline
facilities and for stimulating discussions.
[1] G. Drewes et al., Cell 89 (1997) 297
[2] J. Biernat et al., Neuron 11 (1993) 153
[3] J. Pellettieri and G. Seydoux, Science 298 (2002) 1946
[4] S. Panneerselvam et al., Structure 14 (2006) 173