Protein Technologies

  • Intro
  • Services
  • Equipment
  • Prices
  • Contacts
  • Publications
  • Instruct

Protein Service (PS) offers recombinant protein expression, protein purification and biophysical characterization for drug discovery and life science research. The proteins may be used in structural biology, protein-ligand and protein-protein interaction studies, and in development of diagnostics.

Protein Service is situated in BioMediTech (BMT, Tampere) and the mammalian expression system is operated by Haartman Institute (Helsinki).

The facility offers hands-on counseling concerning protein expression methods and expression vectors, but customer is typically responsible for the preparation of the DNA plasmid. Typical workflow includes pilot scale protein production and purification, scale-up to liter scale, protein isolation and purification and finally, protein characterization by various biochemical and biophysical methods including interaction assays.

Note: Protein Service is capable only work with Biosafety level 1 products.


Order form

Protein Service Order Form including:

  • Research Agreement - Business Operations (University of Tampere)
  • General terms and conditions - Business Operations (University of Tampere)


Protein expression services

Three expression systems (bacteria, insect and mammalian cells) are currently available and they offer potential for large variety of different types of proteins. Potential customer is asked to contact the coordinator of the facility for further information in case the selection of the suitable expression system is not straightforward.

Protein expression in E. coli:

Customer provides expression vector and the expression will be first confirmed utilizing bottle or plate cultures (makes it also possible to optimize growth media). After positive results the production will be performed by using fermentor.

The bioreactor laboratory service offers protein production in bacterial host in 5 l batches using Labfors Infors 3 fermentor. The accurately controlled growth conditions enable higher cell densities and better product quality than by using conventional bottle cultures.

PS sends the final products to the customer and reports SDS-PAGE and Western Blot results (customer is asked to provide antibodies for WB).

Baculovirus Expression Vector System (BEVS):

Customer provides the desired insert in plasmid donor vector and is asked to confirm the clone by DNA sequencing. If necessary, DNA sequencing can be done also in the University of Tampere.

Note! PS kindly asks to measure and label the concentration of the plasmid provided (min 5 ng, V=25 µl).

The donor plasmid is used to prepare six bacmids, which are confirmed by PCR and then three of them are transfected to the Sf9 insect cells (optionally HighFiveTM cells). Volume of secondary virus (P2) is typically 50 ml and expression of protein is confirmed by Western Blot from cell pellet and medium (antibodies against possible purification tag or provided by customer). Virus is stock up for one month and can be used to optimize the expression or for possible production scale-up up to 4.5 L in aliquots of 500 ml. Moreover, protein purification with His-tag or Strept-tag is also possible.

The mammalian cell line for transfection, expression, and large-scale recombinant protein production:

Customer provides the desired cDNA in suitable plasmid vector and is asked to confirm the clone by sequencing. If necessary, sequencing can be done also in the sequencing service of Haartman institute.

Expression vector is transfected to the mammalian cells (CHO-S or HEK 293F) prior to four weeks long cell line selection. SDS-PAGE and Western blot is done from pilot scale expression to confirm protein expression. Cell lines are stored for four months so they can be used to the further expression. Decision of scale-up is asked to give in a month. In scale up cells are first adapted to the suspension growth, and protein expression is confirmed with SDS-PAGE and Western blot from 50 ml patch. This step takes about two weeks, and then scale up to the 5 L is possible.


Protein isolation, purification and characterization services

Host cells are collected by centrifugation and lysed by using either a high-power sonicator or high-pressure homogenizator Emulsiflex. Protein isolation and purification is typically done by chromatographic methods. The protein quality can be evaluated by several methods, including analytical gel filtration, SDS-PAGE, 2-D electrophoresis, dynamic light scattering, calorimetric methods and biosensors. Proteins can be delivered either in solution or as freeze-dried.

Equipment tab provides a brief summary of the methods and instrumentation available.



Chromatography systems. The facility has three complete ÄKTA chromatography systems installed for preparative chromatography work and they all include UV/VIS and conductivity detectors and are equipped with fraction collectors. One of the instruments is equipped with sample pump. We have also recently installed Shimadzu/Malver instrument especially suitable for analytical chromatography. This instrument has gradient pump, autosampler and fraction collector, and is equipped with UV/VIS, fluorescence, DLS and SLS detectors.

Instrument: Shimadzu Prominence 20 HPLC equipped with Malvern microV DLS/SLS detector. The HPLC has autosampler, fluorescence detector, UV/VIS and fraction collector. All the components are made of inert material and the system is installed in refrigerated cabin.


Dynamic light scattering (DLS) allows study of particle size from hydrodynamic diameter of 1 nm (size of sucrose molecule) and particle size distribution (homogeneity) in soluble samples. In addition, it is possible to measure Zeta potential which is a measure of the magnitude of the repulsion or attraction between particles. The significance of zeta potential is that it can be related to the stability of particle dispersions and allows one to find suitable conditions for samples (proteins, virus particles, nanoparticles etc). DLS is thus efficient method both in protein characterization and in protein formulation.

Instrument: Zetasizer Nano ZS (ZEN 3600)


Differential scanning calorimetry (DSC) is primarily used to characterize stability and folding of macromolecules such as proteins. In addition, it can be used to screen protein-ligand interactions, because when a ligand preferentially binds to the native form of a protein, the protein is stabilized and the thermal transition midpoint (Tm) of the protein-ligand complex is higher than that of the protein in the absence of ligand. Therefore, DSC can be used for ligands with ultra-tight binding constants (1020 M-1) that cannot be measured by other methods, as well as a high throughput screening assay for drug discovery (up to 50 samples per day). The DSC instrument is equipped with refrigerated autosampler, enabling large sample sets.

Instrument: VP capillary-DSC (GE Healthcare Life Sciences, Microcal Inc., Northampton, MA, USA)


Isothermal titration calorimetry (ITC) is a thermodynamic technique that directly measures the heat released or absorbed during a biomolecular binding event. Measurement of this heat allows accurate determination of binding constants (KB), reaction stoichiometry (n), enthalpy (ΔH) and entropy (ΔS), thereby providing a complete thermodynamic profile of the molecular interaction in a single experiment. With ITC it is possible to directly measure sub-millimolar to nanomolar binding constants (102 to 109 M-1) and nanomolar to picomolar binding constants (109 to 1012 M-1) by using the competitive binding technique. Moreover, ITC is true in-solution technique and therefore no labeling or immobilization is required.

Instrument: VP-ITC (GE Healthcare Life Sciences, Microcal Inc., Northampton, MA, USA)


Biolayer interferometry (BLI) is technique, which is used to measure real-time, label-free analysis of biomolecular interactions and to provide information on affinity, kinetics and concentration. BLI technology monitors the binding of proteins and other biomolecules to their partners in real time. The binding interaction is continuously monitored by measuring the change in thickness of the protein layer on the biosensor tip. The method has no need to label the protein with fluorescent or chromogenic tags, thus eliminating interference issues. In addition, instrument uses 96 or 384 plates and 16 parallel sensors enabling high throughput assays.

Instrument: Octet® RED384 (Pall ForteBio Corp., Menlo Park, CA, USA)


Surface plasmon resonance (SPR) use an optical method to measure the refractive index near a sensor surface in order to detect an interaction of one molecule that is immobilised onto the sensor’s surface. Method is used e.g. to search for binding partners or most commonly to measure kinetics of an interaction, i.e. the rates of complex formation (ka) and dissociation (kd). The previous parameters can be determined from the information in a sensorgram.

Instrument: Biacore X (GE Healthcare)



Service Academic price
E. coli expression and characterization  
Transformation per plasmid 20 €
Production of a pre-culture from a tested clone 20 €
5 L Bacterial expression culture from pre-culture* 300 €
Affinity Chromatography** 150 €
SDS-PAGE with Coomassie staining 50 €
SDS-PAGE with Oriole staining 70 €
SDS-PAGE with Silver staining 80 €
Immunoblotting*** 100 €
ITC analysis per sample 50 €
ITC equipment usage per day**** 100 €
DSC analysis per sample (Minimum charge 3 samples) 20 €
DLS analysis per sample 20 €
BEVS  
Bacmid production (20 € x 6) 120 €
Transfection Sf9 (HighFive 60 €) 50 €
Optimization of the production by post infection follow up 60 €
Pilot purification assay 40 €
Scale up 1st 500 ml 125 €
Scale up 2nd and following 500 mls 75 €
Mammalian cell expression system  
Transfection and selection 150 €
Cell adaptation 100 €
Scale up 1 L 200 €
Concentration 350 €

Protein Service reserves the right to price adjustments.

* Includes basic reagents and cell collection
** Affinity matrix will be charged separately
*** Customer provides / pays for the antibody
**** User must attain tutoring for usage before starting experiments


Head of the facility:

Vesa Hytönen, Ph.D., Academy Research Fellow
vesa.hytonen(at)uta.fi
Tel: +358 40 190 1517
Room: ARVO F331


Co-ordinator:

Juha Määttä, Ph.D.
juha.maatta(at)uta.fi
Tel: +358 50 318 5814
Room: ARVO F303


Reference publications

Bandaranayake RM, Ungureanu D, Shan Y, Shaw DE, Silvennoinen O, Hubbard SR
Crystal structures of the JAK2 pseudokinase domain and the pathogenic mutant V617F
Nat Struct Mol Biol. 2012 Aug 14;18(9):971-6

Durdagi S, Vullo D, Pan P, Kähkönen N, Määttä JA, Hytönen VP, Scozzafava A, Parkkila S, Supuran CT
Protein-protein interactions: inhibition of mammalian carbonic anhydrases I-XV by the murine inhibitor of carbonic anhydrase and other members of the transferrin family
J Med Chem. 2012 Jun 14;55(11):5529-35

Koho T, Mäntylä T, Laurinmäki P, Huhti L, Butcher SJ, Vesikari T, Kulomaa MS, Hytönen VP
Purification of norovirus-like particles (VLPs) by ion exchange chromatography
J Virol Methods. 2012 Apr;181(1):6-11


Instruct

The modern infrastructure for macromolecular X-ray crystallography and protein characterization is available for European researchers through Instruct, an EU-ESFRI program. Instruct aims at fostering the development of structural biology in Europe. It is coordinated by the Instruct office in Oxford, UK.


Biocenter Finland Structural Biology network formed the framework for an Instruct National Affiliate Center (NAC) established in 2014. The Finnish Instruct-NAC has nodes at the Universities of Helsinki, Oulu, Tampere, Turku, Eastern Finland, and at the Åbo Academi University.

Participation in Instruct through the Instruct-NAC provides access to other European centers of the Instruct network for the researchers in Tampere and it makes the infrastructure and expertise at the BioMediTech institute available for European researchers. Instruct also supports organization of international courses in Tampere.


Structural biology in Tampere

Tampere protein service in collaboration with Helsinki unit offers variety of possibilities to express recombinant proteins for structural biology groups. Both groups in Tampere and Helsinki have a strong background at the manufacturing proteins for structural biology as well as many other purposes for example VLPs and antigens.

In addition, we provide characterization and interaction analysis for the proteins with experimental methods such as ITC, DSC, DLS and OctetRed.

Equipment available in Tampere


Contact persons

Juha Määttä (Tampere, E.Coli and insect cells) juha.maatta[at]uta.fi

Olli Ritvos (Helsinki, mammalian cells) olli.ritvos[at]helsinki.fi