Center for Drug Discovery and Innovation
Protein Production Core
Protein production, a combined CDDI and USF College of Medicine core, is a significant
bottleneck in early phase drug discovery. Two laboratories serve this mission housed
in CDDI and COM under the direction of a core director. The production laboratories
are fully equipped and staffed to help the researcher produce proteins in bacteria,
yeast or insect cells. Whether the protein is hopelessly insoluble or difficult to
produce in quantity the core staff is ready to work with individuals to turn failure
Expression Vector Construction
- We have stocks of expression vectors with different combinations of fusion tags, including polyhistidine-tag (6xHis), Glutathione S-transferase (GST), Maltose-binding protein (MBP), Nus A, and solubility-enhancement tag (SET). We can also use customer-supplied vectors.
- Recombinant proteins would be expressed in bacteria (E. coli), yeast and baculovirus (insect cells) systems. Small-scale trials would be used to find the best condition and cell line for expression and solubility level. Large-scale expression can be performed in New Brunswick BioFlo 310 fermentor (1-14 liter) or temperature-controlled shaking incubators.
- A solubility screening system has been set up to search the best lysis buffer combination for difficult projects. Our latest trial showed about 26% increase in solubility of a target protein.
- The core is equipped with multiple GE AKAT systems and a Bio-Rad Duo Flow system. We provide protein purification using affinity (6xHis, GST, etc), ion-exchange, and/or size exclusion chromatography, from a few to several hundred milligrams protein. The target could be from either natural or recombinant sources.
- The core is equipped with a state-of-the-art nano crystallization robot that can screen thousands of conditions in a matter of hours. It is capable of dispensing without contact a nanoliter quantity of sample both accurately and reproducibly. We have regular synchrotron X-ray access and the ability to perform remote data collection of macromolecule X-ray diffraction.
- We provide regular protein characterization services including SDS-PAGE, western blot
and quantitation of purified protein using NanoDrop. The core is also equipped with
a Bio-Rad Plex 200 (Luminex) XMap Technology system and a Q-Sense E4 Quartz Crystal
Microbalance - with dissipation monitoring (QCM-D).
Major Equipment and Instrumentation
- GE AKTA Explorer (housed in COM)
- GE AKTA Purifier
- Pre-packed Ni-affinity, GST-affinity, Ion-exchange and Size Exclusion Chromatography columns
- Bio-Rad BioLogic Duo Flow FPLC system
- NBS BioFlow 310 Fermentor with 2.5 and 14 liter capacity
- Temperature-controlled Shaking Incubators
- ARI Crystal Phoenix Liquid Handling System (housed in COM)
- Insect cell culture facilities (housed in COM)
- Sorvall RC6 high speed centrifuge with 4x1000 capacity
- French Press and Misonic 4000 sonicator
The core offers both training and production services to our users. We provide theoretical
session and hands-on practice, covering all the services. Once trained, users can
significantly lower their cost and bring the knowledge back to their own labs.
Date and time: Saturday Jan 26, 2013, 10 a.m. - 4 p.m.
Capacity: 8 students / class
Location: Interdisciplinary Research Building (IDRB) room 302 & 319,
3720 Spectrum Blvd.
Tampa, Florida 33612
- Understand the FPLC system and its components
- Learn to plumb and maintain pumps, detectors, loops, columns and other components
- Become familiar with the system software- control the FPLC, write methods and evaluate results
- Gain hands-on laboratory experience by performing a separation, starting with a mixture
of protein standards
Location and Contact Information
Interdisciplinary Research (IDR) Building
3720 Spectrum Blvd., Suite 303
Tampa, Florida 33612
P: (813) 974-0274