Jeff Gelles, Ph.D.
Brandeis University, Aron and Imre Tauber Professor
of Biochemistry and Molecular Pharmacology
Motor Enzymes: Single-molecule Biochemistry
Ph.D., California Institute of Technology
Little Engine Shop
The general interest of our laboratory is macromolecular
motion. Movement is essential for the biological function
of enzymes and nucleic acids. A major focus of our research
is motor enzymes. These remarkable molecular machines catalyze
a chemical reaction, capture the free energy released by
the reaction, and use this energy to perform biologically
useful mechanical work. Motor enzymes play essential roles
in diverse biological processes ranging from muscle contraction
to neuronal development to mitosis to gene transcription.
To learn how the enzymes work, we study them in vitro using molecular cloning, enzymology, protein chemistry, and biophysical chemistry
techniques. The laboratory has also pioneered methods for
visualizing nanometer-scale movements and individual chemical
reaction events in single enzyme molecules. Such methods
reveal the crucial dynamic features of enzyme mechanisms
that are missing from the information produced by static
techniques like x-ray crystallography. In addition, single
molecule methods can examine reaction mechanism steps that
are difficult or impossible to study by conventional biochemical
techniques, which are limited to analyzing the population-average
properties of large molecular ensembles.
The motor enzyme kinesin binds to subcellular organelles and then transports the organelles through the
cytoplasm by pulling them along microtubules. Kinesin or
its homologs drive chromosome movement in meiosis and mitosis,
axonal transport, and formation of the endoplasmic reticulum.
Like other motor enzymes, a kinesin molecule is a chemically-powered
molecular engine: it catalyzes the hydrolysis of ATP and
uses the energy from this reaction to move the enzyme from
site to site on the microtubule lattice. We want to learn
how this engine works by elucidating the steps in the movement
mechanism and determining how these steps are coupled to
the reactions of ATP hydrolysis. A major direction of our
work is to prepare kinesin derivatives using directed mutagenesis
techniques. Once this is done, we analyze the structural
and functional properties of the derivatives. These studies
have demonstrated the importance of particular enzyme structures
for efficient processive movement along the microtubule,
and we are now investigating how these structures function.
One way we study function is to tag single enzyme molecules
with microscopic polystyrene beads and then directly observe
movement along microtubules by light microscopy. Digital
image processing techniques allow us to track bead positions
with nanometer-scale precision, allowing us to directly
observe the kinetics of kinesin-microtubule interactions
and the pattern of single kinesin molecule movements on
the microtubule lattice.
Enzymes that move processively along DNA -- DNA polymerases, RNA polymerases, and DNA helicases
are some examples -- are also motor enzymes. Our laboratory
has developed the first methods for directly observing the
movement of single enzyme molecules along DNA and for measuring
the associated forces. This approach has allowed us to investigate
important mechanistic features of these enzymes that could
not previously be studied. For example, our studies on the E. coli RNA polymerase have revealed the kinetic
behavior of the enzyme at factor-independent transcriptional
terminators. We have also demonstrated that this enzyme
is the most powerful molecular motor yet characterized and
is by itself capable of overcoming mechanical obstacles
to transcription that exist in living cells. The overall
goal of our studies in this area is to explore the translocation
and regulation mechanisms of DNA motor enzymes and, by doing
so, to more completely define the molecular bases for DNA
replication, transcription and recombination.
Please see the The Little Engine Shop page for more information
about the lab, including movies of single-molecule biophysics
Selected recent publications from
- RNA polymerase approaches its promoter without long-range sliding along DNA. Friedman LJ, Mumm JP, Gelles J. Proc Natl Acad Sci U S A. 2013 May 29. PMID: 23720315.
- Single-molecule colocalization FRET evidence that spliceosome activation precedes stable approach of 5' splice site and branch site. Crawford, D.J., Hoskins, A.A., Friedman, L.J., Gelles, J. & Crawford, D.J., Hoskins, A.A., Friedman, L.J., Gelles, J. & Moore, M.J. PNAS (2013). doi:10.1073/pnas.1219305110. [abstract]
- Pathway of actin filament branch formation by Arp2/3 complex revealed by single-molecule imaging. Smith BA, Daugherty-Clarke K, Goode BL, Gelles J. Proc Natl Acad Sci U S A. 2013 Jan 22;110(4):1285-90. doi: 10.1073/pnas.1211164110. Epub 2013 Jan 4. [abstract]
- Operator sequence alters gene expression independently of transcription factor occupancy in bacteria. Garcia HG, Sanchez A, Boedicker JQ, Osborne M, Gelles J, Kondev J, Phillips R. Cell Rep. 2012 Jul 26;2(1):150-61. doi: 10.1016/j.celrep.2012.06.004. Epub 2012 Jul 12 [abstract].
- Rocket launcher mechanism of collaborative actin assembly defined by single-molecule imaging. Breitsprecher D, Jaiswal R, Bombardier JP, Gould CJ, Gelles J, Goode BL. Science. 2012 Jun 1;336(6085):1164-8. doi: 10.1126/science.1218062. [abstract]
- Mechanism of transcription initiation at an activator-dependent promoter defined by single-molecule observation. Friedman LJ, Gelles J. Cell. 2012 Feb 17;148(4):679-89. doi: 10.1016/j.cell.2012.01.018. [abstract]
- New insights into the spliceosome by single molecule fluorescence microscopy. Hoskins AA, Gelles J, Moore MJ. Curr Opin Chem Biol. 2011 Dec;15(6):864-70. doi: 10.1016/j.cbpa.2011.10.010. Epub 2011 Nov 5. Review.
- Mechanism of transcriptional repression at a bacterial promoter by analysis of single molecules. Sanchez A, Osborne ML, Friedman LJ, Kondev J, Gelles J. EMBO J. 2011 Aug 9;30(19):3940-6. doi: 10.1038/emboj.2011.273. [abstract]
- Ordered and dynamic assembly of single spliceosomes. Hoskins AA, Friedman LJ, Gallagher SS, Crawford DJ, Anderson EG, Wombacher R, Ramirez N, Cornish VW, Gelles J, Moore MJ. Science. 2011 Mar 11;331(6022):1289-95. doi: 10.1126/science.1198830. [abstract]
View Complete Publication List on PubMed: Jeff
Last review: June 4, 2013