Kimmo Pääkkönen

Structural insight into dimeric interaction of the SARAH domains from Mst1 and RASSF family proteins in the apoptosis pathway

In eukaryotic cells, apoptosis and cell cycle arrest by the Ras → RASSF → MST pathway are controlled by the interaction of SARAH (for Salvador/Rassf/Hippo) domains in the C-terminal part of tumor suppressor proteins. The Mst1 SARAH domain interacts with its homologous domain of Rassf1 and Rassf5 (also known as Nore1) by forming a heterodimer that mediates the apoptosis process. Here, we describe the homodimeric structure of the human Mst1 SARAH domain and its heterotypic interaction with the Rassf5 and Salvador (Sav) SARAH domain. The Mst1 SARAH structure forms a homodimer containing two helices per monomer. An antiparallel arrangement of the long α-helices (h2/h2′) provides an elongated binding interface between the two monomers, and the short 310 helices (h1/h1′) are folded toward that of the other monomer. Chemical shift perturbation experiments identified an elongated, tight-binding interface with the Rassf5 SARAH domain and a 1:1 heterodimer formation. The linker region between the kinase and the SARAH domain is shown to be disordered in the free protein. These results imply a novel mode of interaction with RASSF family proteins and provide insight into the mechanism of apoptosis control by the SARAH domain.

Quaternary structure built from subunits combining NMR and small-angle x-ray scattering data.

A new principle in constructing molecular complexes from the known high-resolution domain structures joining data from NMR and small-angle x-ray scattering (SAXS) measurements is described. Structure of calmodulin in complex with trifluoperazine was built from N- and C-terminal domains oriented based on residual dipolar couplings measured by NMR in a dilute liquid crystal, and the overall shape of the complex was derived from SAXS data. The residual dipolar coupling data serves to reduce angular degrees of freedom, and the small-angle scattering data serves to confine the translational degrees of freedom. The complex built by this method was found to be consistent with the known crystal structure. The study demonstrates how approximate tertiary structures of modular proteins or quaternary structures composed of subunits can be assembled from high-resolution structures of domains or subunits using mutually complementary NMR and SAXS data.

The physical character of information

The mathematical theory of communication defines information in syntax without reference to its physical representation and semantic significance. However, in an everyday context, information is tied to its representation and its content is valued. The dichotomy between the formal definition and the practical perception of information is examined by the second law of thermodynamics that was recently formulated as an equation of motion. Thermodynamic entropy shows that the physical representation of information is not inconsequential in generation, transmission and processing of information. According to the principle of increasing entropy, communication by dissipative transformations is a natural process among many other evolutionary phenomena that level energy-density differences between components of a communication system and its surroundings. In addition, information-guided processes direct down along descents on free energy landscapes. The non-integrable equation for irreversible processes reveals that there is no universal analytical algorithm to match source to channel. Noise infiltration is also regarded by the second law as an inevitable consequence of energy transduction between a communication system and its surroundings. Communication is invariably associated with misunderstanding because mechanisms and means of information processing at the receiver differ from those at the sender. The significance of information is ascribed to the increase in thermodynamic entropy in the receiver system that results from execution of the received message.

Structural basis of PxxDY motif recognition in SH3 binding.

We have determined the solution structure of epidermal growth factor receptor pathway substrate 8 (Eps8) L1 Src homology 3 (SH3) domain in complex with the PPVPNPDYEPIR peptide from the CD3epsilon cytoplasmic tail. Our structure reveals the distinct structural features that account for the unusual specificity of the Eps8 family SH3 domains for ligands containing a PxxDY motif instead of canonical PxxP ligands. The CD3epsilon peptide binds Eps8L1 SH3 in a class II orientation, but neither adopts a polyproline II helical conformation nor engages the first proline-binding pocket of the SH3 ligand binding interface. Ile531 of Eps8L1 SH3, instead of Tyr or Phe residues typically found in this position in SH3 domains, renders this hydrophobic pocket smaller and nonoptimal for binding to conventional PxxP peptides. A positively charged arginine at position 512 in the n-Src loop of Eps8L1 SH3 plays a key role in PxxDY motif recognition by forming a salt bridge to D7 of the CD3epsilon peptide. In addition, our structural model suggests a hydrogen bond between the hydroxyl group of the aromatic ring of Y8 and the carboxyl group of E496, thus explaining the critical role of the PxxDY motif tyrosine residue in binding to Eps8 family SH3. These finding have direct implications also for understanding the atypical binding specificity of the amino-terminal SH3 of the Nck family proteins.

Solution structures of the first and fourth TSR domains of F-spondin

F‐spondin is a protein mainly associated with neuronal development. It attaches to the extracellular matrix and acts in the axon guidance of the developing nervous system. F‐spondin consists of eight domains, six of which are TSR domains. The TSR domain family binds a wide range of targets. Here we present the NMR solution structures of TSR1 and TSR4. TSR domains have an unusual fold that is characterized by a long, nonglobular shape, consisting of two β‐strands and one irregular extended strand. Three disulfide bridges and stack of alternating tryptophan and arginine side‐chains stabilize the structure. TSR1 and TSR4 structures are similar to each other and to the previously determined TSR domain X‐ray structures from another protein, TSP, although TSR4 exhibits a mobile loop not seen in other structures. Proteins 2006.

Solution structure of the GUCT domain from human RNA helicase II/Guβ reveals the RRM fold, but implausible RNA interactions

Human RNA helicase II/Guα (RH‐II/Guα) and RNA helicase II/Guβ (RH‐II/Guβ) are paralogues that share the same domain structure, consisting of the DEAD box helicase domain (DEAD), the helicase conserved C‐terminal domain (helicase_C), and the GUCT domain. The N‐terminal regions of the RH‐II/Gu proteins, including the DEAD domain and the helicase_C domain, unwind double‐stranded RNAs. The C‐terminal tail of RH‐II/Guα, which follows the GUCT domain, folds a single RNA strand, while that of RH‐II/Guβ does not, and the GUCT domain is not essential for either the RNA helicase or foldase activity. Thus, little is known about the GUCT domain. In this study, we have determined the solution structure of the RH‐II/Guβ GUCT domain. Structural calculations using NOE‐based distance restraints and residual dipolar coupling‐based angular restraints yielded a well‐defined structure with β‐α‐α‐β‐β‐α‐β topology in the region for K585‐A659, while the Pfam HMM algorithm defined the GUCT domain as G571‐E666. This structure‐based domain boundary revealed false positives in the sequence homologue search using the HMM definition. A structural homology search revealed that the GUCT domain has the RRM fold, which is typically found in RNA‐interacting proteins. However, it lacks the surface‐exposed aromatic residues and basic residues on the β‐sheet that are important for the RNA recognition in the canonical RRM domains. In addition, the overall surface of the GUCT domain is fairly acidic, and thus the GUCT domain is unlikely to interact with RNA molecules. Instead, it may interact with proteins via its hydrophobic surface around the surface‐exposed tryptophan. Proteins 2009.

Conformations of the Regulatory Domain of Cardiac Troponin C Examined by Residual Dipolar Couplings; Complex with peptide

Conformations of the regulatory domain of cardiac troponin C (cNTnC) were studied by means of residual dipolar couplings measured from samples dissolved in dilute liquid crystals. Changes in the main chain HN residual dipolar couplings revealed a conformational change in cNTnC due to the complexation with the second binding region (amino acids 148-163) of cardiac troponin I (cTnI). Formation of the complex is accompanied with a molecular realignment in the liquid crystal. The residual dipolar couplings measured for apo-cNTnC and the complex with TnI were in agreement with the values computed from the corresponding closed and open solution structures, whereas for the calcium-loaded conformation the correlation and quality factor were only modest. Ca2+-cNTnC may be subject to conformational exchange. The data support the model that cardiac troponin C functions as a calcium-dependent open-closed switch, such as the skeletal troponin C.