Deuterium Labeling Together with Contrast Variation Small-Angle Neutron Scattering Suggests How Skp Captures and Releases Unfolded Outer Membrane Proteins

Published

August 4, 2016

Author(s)

Nathan R. Zaccai, Clifford W. Sandlin, James T. Hoopes, Joseph E. Curtis, Patrick J. Fleming, Karen G. Fleming, Susan T. Krueger

Abstract

In gram¿negative bacteria, the chaperone protein Skp forms specific and stable complexes with membrane proteins as these are transported across the periplasm to the outer membrane. The jellyfish¿like architecture of Skp is similar to the eukaryotic and archeal prefoldins and the mitochondrial TIM chaperones. α¿helical 'tentacles¿ extend from a β¿strand 'body¿ to create an internal cavity. Contrast variation small¿angle neutron scattering (SANS) experiments on Skp alone in solution and bound to two unfolded outer membrane proteins (uOMPs), OmpA and OmpW, demonstrated that the helical tentacles of Skp bind their substrate in a clamp¿like mechanism, in a conformation similar to that previously observed in the apo crystal structure of Skp. Deuteration of the uOMP component allowed the shapes of Skp and uOMP, as well as the location of uOMP with respect to Skp to be determined in both complexes. The data yield the first direct structural evidence that the α¿helical Skp tentacles move close together on binding its substrate and that the structure of Skp is different when binding different uOMPs. This represents unique information that could not be obtained without deuterium labeling of the uOMPs. The SANS analysis provides experimental support for a simple clamp¿like mechanism used by jellyfish¿like chaperones.

Citation

Methods in Enzymology

Volume

566

Publisher Info

Elsevier, Oxford, -1

Pub Type

Books

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Keywords

Skp, jellyfish-like chaperone, holdase, periplasm, Outer Membrane Protein (OMP) transport, disordered protein, small-angle neutron scattering, SANS

Citation

Zaccai, N. , Sandlin, C. , Hoopes, J. , Curtis, J. , Fleming, P. , Fleming, K. and Krueger, S. (2016), Deuterium Labeling Together with Contrast Variation Small-Angle Neutron Scattering Suggests How Skp Captures and Releases Unfolded Outer Membrane Proteins, Elsevier, Oxford, -1, [online], https://tsapps.nist.gov/publication/get_pdf.cfm?pub_id=918519 (Accessed December 13, 2024)