Markku S. Kulomaa, Professor
Institute of Medical Technology (IMT)
Biokatu 6
FI-33014 University of Tampere
Phone: +358-3-3551-8499
Fax: +358-3-3551 7710
Graduate students:
Tiia Koho
Soili Lehtonen

Protein engineering and protein design are the major experimental approaches in the molecular biotechnology (MBT) group in IMT. In order to study, at a molecular level, protein structure-function relationship, the proteins are produced as recombinant forms and modified by site-directed mutagenesis. Bacterial (E. coli) and eukaryotic (baculovirus and insect cells) production systems and a phage display system are used to produce the re-proteins. Purification of recombinant proteins is done by chromatographic methods, particularly by affinity chromatography. The functional characterization of protein-ligand and protein-protein interactions is performed using a wide variety of chemical and biophysical methods. They include for instance use of surface plasmon resonance (SPR) or optical biosensors, fluorescence spectroscopy, isothermal titration calorimetry (ITC), differential scanning calorimetry (DSC) and atomic force microscopy (AFM).

In addition to fundamental structure-function research of interaction between avidin (Avd) and biotin, avidin-biotin technology is a major objective in the research projects including applications from life science and medicine to nanoscience and nanotechnology. In addition to novel protein tools, new modified ligands have also been produced and characterized for improvement and development of novel approaches in avidin-biotin technology. In one of our recent innovations, circularly permuted subunit fusions were used to create "dual chain" (dcAvd) and "single chain" (scAvd) Avds which have two and four biotin-binding sites in a polypeptide chain, respectively. They have provided multipurpose scaffolds which can be used to generate valuable protein molecules for scientific and applicative purposes. In contrast to wt Avd (four identical subunits), scAvd enables each of the four domains to be independently modified. The dcAvd and scAvd scaffold can thus be used to construct spatially and stoichiometrically defined pseudo-tetrameric Avds with altered domain characteristics. This subunit-to-domain strategy should also be applicable to other proteins and protein complexes in new applications in life sciences and nanotechnology. The avidin research is performed as an active collaborative project with several research groups from Finland and abroad.