Dataset Information

The high-resolution NMR structure of the early folding intermediate of the Thermus thermophilus ribonuclease H.

ABSTRACT: Elucidation of the high-resolution structures of folding intermediates is a necessary but difficult step toward the ultimate understanding of the mechanism of protein folding. Here, using hydrogen-exchange-directed protein engineering, we populated the folding intermediate of the Thermus thermophilus ribonuclease H, which forms before the rate-limiting transition state, by removing the unfolded regions of the intermediate, including an alpha-helix and two beta-strands (51 folded residues). Using multidimensional NMR, we solved the structure of this intermediate mimic to an atomic resolution (backbone rmsd, 0.51 A). It has a native-like backbone topology and shows some local deviations from the native structure, revealing that the structure of the folded region of an early folding intermediate can be as well defined as the native structure. The topological parameters calculated from the structures of the intermediate mimic and the native state predict that the intermediate should fold on a millisecond time scale or less and form much faster than the native state. Other factors that may lead to the slow folding of the native state and the accumulation of the intermediate before the rate-limiting transition state are also discussed.

SUBMITTER: Zhou Z

PROVIDER: S-EPMC2628467 | biostudies-literature | 2008 Dec

REPOSITORIES: biostudies-literature

ACCESS DATA

Publications

The high-resolution NMR structure of the early folding intermediate of the Thermus thermophilus ribonuclease H.

Zhou Zheng Z Feng Hanqiao H Ghirlando Rodolfo R Bai Yawen Y

Journal of molecular biology 20080926 2

Elucidation of the high-resolution structures of folding intermediates is a necessary but difficult step toward the ultimate understanding of the mechanism of protein folding. Here, using hydrogen-exchange-directed protein engineering, we populated the folding intermediate of the Thermus thermophilus ribonuclease H, which forms before the rate-limiting transition state, by removing the unfolded regions of the intermediate, including an alpha-helix and two beta-strands (51 folded residues). Using ...[more]

PMID: 18848567

Similar Datasets

Project description:The fundamental chemistry underpinning aerobic life on Earth involves reduction of dioxygen to water with concomitant proton translocation. This process is catalyzed by members of the heme-copper oxidase (HCO) superfamily. Despite the availability of crystal structures for all types of HCO, the mode of action for this enzyme is not understood at the atomic level, namely how vectorial H(+) and e(-) transport are coupled. Toward addressing this problem, we report wild type and A120F mutant structures of the ba(3)-type cytochrome c oxidase from Thermus thermophilus at 1.8 Å resolution. The enzyme has been crystallized from the lipidic cubic phase, which mimics the biological membrane environment. The structures reveal 20 ordered lipid molecules that occupy binding sites on the protein surface or mediate crystal packing interfaces. The interior of the protein encloses 53 water molecules, including 3 trapped in the designated K-path of proton transfer and 8 in a cluster seen also in A-type enzymes that likely functions in egress of product water and proton translocation. The hydrophobic O(2)-uptake channel, connecting the active site to the lipid bilayer, contains a single water molecule nearest the Cu(B) atom but otherwise exhibits no residual electron density. The active site contains strong electron density for a pair of bonded atoms bridging the heme Fe(a3) and Cu(B) atoms that is best modeled as peroxide. The structure of ba(3)-oxidase reveals new information about the positioning of the enzyme within the membrane and the nature of its interactions with lipid molecules. The atomic resolution details provide insight into the mechanisms of electron transfer, oxygen diffusion into the active site, reduction of oxygen to water, and pumping of protons across the membrane. The development of a robust system for production of ba(3)-oxidase crystals diffracting to high resolution, together with an established expression system for generating mutants, opens the door for systematic structure-function studies.

Dataset Information

The high-resolution NMR structure of the early folding intermediate of the Thermus thermophilus ribonuclease H.

Publications

The high-resolution NMR structure of the early folding intermediate of the Thermus thermophilus ribonuclease H.

Similar Datasets

OmicsDI is part of the ELIXIR infrastructure

Tweets