Core Facilities

carlo-carolis
Biomolecular Screening and Protein Technologies Unit

Unit Structure

HEAD OF THE UNIT:
Carlo Carolis
TECHNICAL OFFICERS:
Miriam Alloza, Silvia Speroni
VISITING SCIENTIST:
Melissa Rieger (June – December 2013)
TRAINEE:
Judit García González (June – July 2013)

Summary

The Biomolecular Screening & Protein Technologies Facility is a shared resource facility accessible to CRG and non-CRG scientists.

The Facility offers the following services:

1. Recombinant plasmid DNA engineering
2. Analytical and preparative scale expression of nascent or epitope-tagged recombinant proteins
3. Protein purification
4. Biophysical analysis of molecular interactions
5. Automation technology for screening assays

These goals are accomplished by a centralized laboratory with dedicated, experienced staff, which enables high-throughput, economy of scale, virus preparation and protein expression services, including quality assurance and control procedures to ensure efficient, consistent production and purification of recombinant proteins. Many recombinant proteins produced by the facility have been used for analytical biochemistry studies designed to investigate enzymatic properties, structure-function relationships between protein-protein, protein-nucleic-acid, and protein-small molecule interactions, custom antibody production.

In addition, the mission of the facility is to enable researchers to apply cutting edge technology and unique resources to discover molecular, genetic, and small molecule compounds suitable to further study the functions of poorly understood proteins, signalling pathways, and cells in complex biological processes.


Highlights

  • A new method for DNA assembly was implemented in the Unit, reducing the cost of standard cloning procedures, and allowing inserting specific ORFs in several expression vectors in a shorter time. This method was disseminated to the entire PRBB by organizing specific courses in collaboration with people from the Guigó and Serrano labs. The facility has been engaged in the preparation of a new mix for restriction free cloning strategy.  More than 1,000 aliquots have been distributed among 12 CRG laboratories.
  • The unit has worked on increasing the efficiency and throughput in protein production, and significantly reducing the lead-time for the identification of soluble proteins. For instance, new methods have been implemented in the 2 ÄKTAxpress™ liquid chromatography instruments in order to speed up the procedure for antibody purification and improving 2-step automated purification protocols. With the implementation of this method we are able to run 8 parallel protein purification experiments overnight.
  • A NanoTemper´s Monolith platform was installed in the facility. It utilizes Microscale Thermophoresis to enable real-time, immobilization free analysis of biomolecules providing information on the affinity, stoichiometry and aggregation properties of biomolecules in buffers and complex biological liquids including blood serum and cell lysate. This is the first system of the sort installed in Spain.
  • A transient mammalian expression system and the baculovirus expression system were inserted in our list of services provided.

 


Selected Publications

  1. Paetzold B, Carolis C, Ferrar T, Serrano L, Lluch-Senar M.
    “In Situ Overlap and Sequence Synthesis During DNA Assembly.”
    ACS Synth Biol, 2(12):750-755 (2013).
  2. Dos Santos HG, Abia D, Janowski R, Mortuza G, Bertero MG, Boutin M, Guarín N, Méndez-Giraldez R, Nuñez A, Pedrero JG, Redondo P, Sanz M, Speroni S, Teichert F, Bruix M, Carazo JM, Gonzalez C, Reina J, Valpuesta JM, Vernos I, Zabala JC, Montoya G, Coll M, Bastolla U, Serrano L.
    “Structure and non-structure of centrosomal proteins.”
    PLoS One, 8(5): e62633 (2013)
  3. Mitrovic S, Nogueira C, Cantero-Recasens G, Kiefer K, Fernández-Fernández JM, Popoff JF, Casano L, Bard FA, Gomez R, Valverde MA, Malhotra V.
    “TRPM5-mediated calcium uptake regulates mucin secretion from human colon goblet cells.”
    Elife, 2:e00658 (2013).