The doctoral dissertations of the former Helsinki University of Technology (TKK) and Aalto University Schools of Technology (CHEM, ELEC, ENG, SCI) published in electronic format are available in the electronic publications archive of Aalto University - Aaltodoc.
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Computational Analysis of Protein Function - Lipases and A-Kinase Anchoring Proteins

Sanna Herrgård

Dissertation for the degree of Doctor of Science in Technology to be presented with due permission of the Department of Chemical Technology for public examination and debate in Auditorium KE 2 (Komppa Auditorium) at Helsinki University of Technology (Espoo, Finland) on the 16th of March, 2001, at 12 noon.

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Abstract

In this thesis, normal mode analysis, electrostatic calculations, and sequence-based bioinformatics methods were applied to investigate protein sequence-structure-function relationship. Two different protein families were studied: a family of fungal lipases (the Rhizomucor miehei lipase or RmL family), and a family of adapter proteins called A-kinase anchoring proteins (AKAPs). The RmL family is well conserved, and three-dimensional structures for several members of this family are known. The AKAP family is a functionally, but not structurally, related protein family that consists of multi-domain proteins whose domain architectures are not well understood.

In the first part of this thesis, lipase structure-function relationship was investigated using two complementary methods: normal mode analysis and electrostatic calculations. Normal mode analysis was applied to characterize the collective motions in RmL; a global breathing motion as well as local loop motions were shown to be associated with the conformational change. The electrostatic calculations yield a detailed description of the role of electrostatic interactions in the RmL structure, function, and stability. Two main results were obtained in this study. First, the key residues that affect the lid stability were identified. Second, a network of electrostatic interactions was discovered. This network is an important feature of the lipase structure, since it connects the active site to the mobile lid region. The network was observed to be conserved in the RmL family and their homologues.

In the second part of the thesis, the sequence-function relationship of an AKAP from Caenorhabditis elegans (AKAPCE) was studied using sequence-based bioinformatics methods. The domain architecture of AKAPCE reveals that it shares two domains with SARA (Smad anchor for receptor activation), which is an adapter protein involved in the TGFβ signaling pathway. One of the domains, FYVE, is well characterized, whereas the other one, a TGFβ receptor binding domain, has not been characterized earlier. The existence of these two domains in AKAPCE leads us to propose a novel AKAP function as a TGFβ receptor binding protein.

This thesis consists of an overview and of the following 4 publications:

  1. S. Jääskeläinen, C.S. Verma, R.E. Hubbard, P. Linko, and L.S.D. Caves. Conformational change in the activation of lipase: an analysis in terms of low-frequency normal modes. Protein Sci., 7:1359-1367, 1998. © 1998 The Protein Society. By permission.
  2. S. Jääskeläinen, C.S. Verma, R.E. Hubbard, and L.S.D. Caves. Identifying key electrostatic interactions in Rhizomucor miehei lipase: the influence of solvent dielectric. Theor. Chem. Acc., 101:175-179, 1999. © 1999 Springer-Verlag. By permission.
  3. S. Herrgard, C.J. Gibas, and S. Subramaniam. Role of an electrostatic network of residues in the enzymatic action of the Rhizomucor miehei lipase family. Biochemistry, 39:2921-293O, 2000. © 2000 American Chemical Society. By permission.
  4. S. Herrgård, P. Jambeck, S.S. Taylor, and S. Subramaniam. Domain architecture of a Caenorhabditis elegans AKAP suggests a novel AKAP function. FEBS Lett., 486:107-111, 2000. © 2000 Elsevier Science. By permission.

Keywords: A-kinase anchoring protein, domain architecture, domain motion, electrostatics, normal mode analysis, Rhizomucor miehei lipase

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© 2001 Helsinki University of Technology

Last update 2011-05-26