Universitetet i Tromsø - Startsida Universitetet i Tromsø - Startsida
Fakturaadresse
UiT Norges arktiske universitet
Fakturamottak
Postboks 6050 Langnes
NO-9037 Tromsø


Organisasjonsnummer
970 422 528

Facilities

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 FacilProdLine

 NORSTRUCT offers an efficient pipeline from recombinant protein production, via preliminary characterization, to crystal structure determination.

The protein expression facility
The protein purification facility
The X-ray crystallography facility
Other facilities

Most of the internal projects are passed through this "production line", starting with target selection using various bioinformatics tools, cloning, expression and fermentation both in small and larger scale. Targets where there is no 3-dimensional structure of homologus proteins are subjected to production using Selenio-methionine (Se-Met) for potential heavy atom phasing.

The lab is well equipped with several liquid chromatography systems for protein purification.

Protein samples are subjected to dynamic light scattering (DLS)analysis prior to crystallisation trials in order to estimate the homogenity of the sample. Circular Dicroism and NMR facilities are available at the Department of Chemistry and MS-MALDI is available at Department of Pharmacy.

Automatic setup of crystallisation experiments is possible using an Impax crystallisation robot.

X-ray data are collected locally using an image detector system with a rotating anode generator. The lab is also frequent users of synchrotron facilities in Europe (ESRF, Bessy, SLS).

Computing facilities for structure determination and structure-function relations include a network of PCs running on linux and windows. A couple of Silicon Graphics (SGI) workstations are also available. Protein structures are routinely solved by molecular replacement, MAD and SAD methods.

Other facilities at NorStruct include microcalorimetry and biocomputing.
 


Protein expression facility
FacilSIXFORS

Cloning

DNA cloning is generally performed using the Gateway Technology system, based on site-specific recombination properties of bacteriophage lambda. The DNA sequences can subsequentially be rapidly moved into multiple vector systems for functional analysis and protein expression.

 FaciFermExpression

The lab has equipment for culturing bacterial and fungal cells, primarily E.coli and P.pastoris, in small and large scale.
Small scale fermentation and optimization is possible using an Infors HT Sixfors bioreactor (6 x 500 ml), while larger volumes (up to 10 l) are produced in the Infors Techfors S 15 L TV fermentor. Both fermentors are capable of automatically regulating temperature, pH and oxygen. The Techfors is also able to automatically control the flow of oxygen.
Proteins are routinely produced with N- or C-terminal tags, such as His, GST and MBP. Proteins where no structure exists of hologouos proteins are produced with Selenio-methionine (Se-Met) in order to overcome the crystallographic "phase problem".

Purification facility

FPLC  The purification lab is well equipped with chromatographic systems from GE Healthcare:

  • Akta Explorer
  • Akta Basic
  • Akta FPLC
  • Akta Prime

    The lab is also equipped with a range of columns for both small and great scale protein purification as well as separation material for the most commonly used separation techniques, as affinity, ion exchange, HIC and size exclusion.

  • X-ray crystallography facility
      

     FacilRobot

     

     

     

     

     

     

     

     

     Pre-crystallisation analysis

    Dynamic light scattering (DLS)
    Protein samples are routinely checked for homogenity prior to crystallisation using dynamic light scattering; DynaPro from Proterion Corporation Ltd. UK.

     Crystallisation robotics

    The lab is equipped with an Douglas Instruments IMPAX I-5 Crystallisation robot. Facilities for variable temperature storage are available.
    An experimental setup for crystallisation experiments requiring inert atmosphere (eg. nitrogen atmosphere for proteins in reduced state) is also available.

     FacilRaxis

     Detector system
    A new Rigaku MicroMax-007HF Microfocus rotating anode generator with Osmic Confocal Max-Flux® High Resolution optic will be installed during the summer of 2006, providing a high brilliance and highly focussed X-ray beam. Data collection at cryogenic temperatures will be enabeled using Rigaku X-stream-2000CE, which operates on compressed air, and thus does not require liquid nitrogen.
    Diffraction data is acquired using an Raxis VI++ image plate detector. Data is processed using the graphical interface CrystalClear or any crystallographic software of the users choice.


     

     FacilESRF

     

     

     

     

     

     

     

     

     

     Synchrotron
    The lab routinely collects X-ray data at synchrotron facilities such as the European Synchrotron Facility (ESRF) in Grenoble, Bessy in Berlin, and Swiss Light Source (SLS) in Zürich.

     


    Other facilities
     FacilBOB Microcalorimetry
    The laboratory is equipped with a differential scanning calorimeter (DSC) and a isothermal titration calorimeter (ITC). These are routinely used to investigated not only the thermodynamic properties of macromolecules and interactions with other molecules but also to explore the kinetics of reactions catalyzed by enzymes.
     FacilComput Biocomputing
    Silicon graphics and Linux workstations are used to investigate experimental and homology modeled atomic structures of macromolecules. More sophisticated computational analysis of macromolecules and their complexes are carried out using resources at the High Performance Computing facilities located at the University campus. Such investigations include molecular dynamics simulations, free energy calculations, scoring and docking problems, protein flexibility, just to mention a few examples.

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    Sist endret: 08.10.2009


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    UiT Norges arktiske universitet - UiT The Arctic University of Norway - UiT Norgga árktalaš universitehta. N-9019 Tromsø - Tlf.: +47 77 64 40 00 - E-post: postmottak@uit.no