NEWS OF THE WEEK SCIENCE
HUGE PROTEIN ANALYZED BY NMR Ability to study large NMR-accessible biomolecules skyrockets
I
NCREASINGLY SOPHISTICATED
nuclear magnetic resonance spectroscopy (NMR) techniques have made it possible to analyze larger and larger biomolecules in the past few years. But structures exceeding 100 kilodaltons have remained extremely difficult or impossible to study by NMR. Now researchers have made nearly an order of magnitude leap in the mass range of proteins amenable to N M R analysis by analyzing the GroEL-GroES chaperone system, a huge 9 0 0 kDa protein complex. Chaperones help proteins fold into thennative states. The analysis was carried out by molecular biophysics professor Kurt Wuthrich and postdoc Jocelyne Fiaux at the Swiss Federal Institute of Technology (ETH), Zurich, in collaboration with genetics professor and Howard Hughes Medical Institute investigator Arthur L. Horwich and postdoc Eric B. Bertelsen at Yale University School of Medicine [Nature, 418,207 (2002)]. They accomplished the feat by combining the principles of two high-mass N M R techniques developed earlier by Wiithrich's group: transverse relaxation-optimized spectroscopy (TROSY) and cross-correlated relaxationenhanced polarization transfer. "The sound barrier has been shattered big-time in terms of what can be looked at with NMR," Horwich says. "This initial study is more proof of principle, but it suggests we will be able to analyze the {GroEL-GroES} system in a way that's never been possible before." HTTP://PUBS.ACS.ORG/CEN
W u t h r i c h , Horwich, and coworkers did not determine the N M R solution structure of GroEL-GroES. Instead, they observed resonances that serve as markers for a significant number of individual protons in the complex, which is much less difficult. But these markers do make it possible to study the protein's local dynamic and structural properties—such as the responses of specific amino acid residues when GroES binds to GroEL. (The major structural breakthrough on GroEL-GroES was actually achieved in 1997, when Horwich's group, along with that of the late Yale biochemistry professor Paul B. Sigler, nailed down the complex's structure using Xray crystallography) The GroES-GroEL system is a favorable case for extremely high mass N M R analysis. GroES has seven identical subunits and GroEL has 14, so the number of unique N M R resonances in GroEL-GroES is much lower— and the effective concentration of each subunit is much higher— than would be the case for a monomelic complex of similar mass. Nevertheless, "the fact that they can study a system of that size is really fantastic," says N I H biophysical chemist Ad Bax. "It potentially brings a whole lot of problems within NMR's reach that we would not have dared to dream about prior to this work. With ongoing improvements in N M R hardware and sensitivity it is quite conceivable that similar data could eventually be obtained for monomelic systems." The development of TROSY "enabled studies of systems up to
CHAPERONE In the GroELGroES complex (shown here in surface and cross-sectional representations), GroES (white) and both halves of GroEL (multicolored and gold) are each made up of seven identical subunits (highlighted in multicolored segment of GroEL). the 100-kDa range," comments chemistry professor David E. ^Jfemmer of the University of California, Berkeley but most people in the field did not believe until now "that studies of still bigger complexes were really possible." Chemistry and biochemistry professor Lewis E. Kay ofthe University of Toronto says that the ability "to get some information about such a huge system is impressive. But we don't have the techniques to be able to assign all the nuclei in a biomolecule that's this large. This study beautifully illustrates the potential for dissecting out very large systems based on having a priori information on the smaller components that go into them."-STU B0RMAN
CHAPERONE TEAM Fiaux (left in photo at bottom) and Wuthrich, and Bertelsen (left in left photo) and Horwich.
C&EN
/ JULY
15, 2002
9
