People - [ Faculty ]
Nearly all of life's essential chemistry is catalyzed by proteins. Each protein is exquisitely tuned for a specific function, from DNA replication to metabolism. Over the past 20 years, scientists have grown to appreciate the importance of dynamics and flexibility in enzyme catalysis. We are interested in understanding the relationship between a protein's flexibility and its function, and in particular how proteins can use structural disorder to their advantage during catalysis.
One system of interest is the Integrase protein (IN) from HIV-1. IN is responsible for inserting the reverse-transcribed viral cDNA into the host's genome. As such, it is an attractive drug target, and pharmaceutical companies are actively working on second generation IN inhibitors as part of a comprehensive AIDS treatmen regimen. The active site of IN is dynamic in the absence of target DNA, yet it adopts a rigid conformation during DNA strand transfer. We are interested in how this disorder to order transition may affect IN specificity and mechanism.
We are also studying the mechanism of the Pin1 peptidyl-prolyl isomerase. Pin1 targets proline residues that follow a phosphoserine (pSer) or phosphothreonine (pThr), and it is thought to play an important role in the cell cycle. Pin1 is a multidomain protein, and its two domains work together to target pSer-Pro and pThr-Pro sequences. Similar to IN, Pin1 undergoes a disorder to order transition upon binding a substrate. We are currently trying to understand how Pin1 reorganizes upon binding and what effect, if any, this transition has on its substrate. Students in the Fitzkee lab use a combination of biochemical approaches along with NMR spectroscopy to study protein structure and dynamics. While we are primarily an experimental group, opportunities often arise to incorporate modeling into our research to interpret the data we collect.
The IN Target Capture Complex. The IN from prototype foamy virus (blue) along with the viral DNA (yellow) and the host DNA (cyan). The structure is poised for strand transfer. Adapted from PDB ID 3OS1 (original structure by Maertens G.N., Hare, S., and Cherepanov, P.).
1: Sgourakis NG, Lange OF, DiMaio F, André I, Fitzkee NC, Rossi P, Montelione GT, Bax A, Baker D. Determination of the structures of symmetric protein oligomers from NMR chemical shifts and residual
dipolar couplings. J Am Chem Soc. 2011 Apr 27;133(16):6288-98.
2:Fitzkee NC, Torchia DA, Bax A. Measuring rapid hydrogen exchange in the homodimeric 36 kDa HIV-1 integrase catalytic core domain. Protein Sci. 2011 Mar;20(3):500-12.
3: Fitzkee NC, Bax A. Facile measurement of ¹H-¹5N residual dipolar couplings in larger perdeuterated
proteins. J Biomol NMR. 2010 Oct;48(2):65-70.
4: Fitzkee NC, Masse JE, Shen Y, Davies DR, Bax A. Solution conformation and dynamics of the
HIV-1 integrase core domain. J Biol Chem. 2010 Jun 4;285(23):18072-84.
5: Fitzkee NC, García-Moreno E B. Electrostatic effects in unfolded staphylococcal nuclease. Protein Sci. 2008 Feb;17(2):216-27.
6: Street TO, Fitzkee NC, Perskie LL, Rose GD. Physical-chemical determinants of turn conformations in globular proteins. Protein Sci. 2007 Aug;16(8):1720-7.
7: Fitzkee NC, Rose GD. Sterics and solvation winnow accessible conformational space for unfolded proteins. J Mol Biol. 2005 Nov 4;353(4):873-87.
8: Fitzkee NC, Fleming PJ, Gong H, Panasik N Jr, Street TO, Rose GD. Are proteins made from a limited parts list? Trends Biochem Sci. 2005 Feb;30(2):73-80.