Interaction of GroEL and GroEL/GroES complexes with a nonnative subtilisin variant: a small-angle neutron scattering study.
Small-angle neutron scattering and contrast variation were used to study the solution structure of GroEL and GroEL/GroES chaperonins complexed with a nonnative variant of the polypeptide substrate, subtilisin (PJ9). The subtilisin was 86% deuterated (dPJ9) so that it contrasted sufficiently with the chaperonin, allowing the contrast variation technique to be used to separate the scattering from the two components bound in the complex. Both the native double-ring GroEL and a single-ring mutant were used with dPJ9 bound in a 1:1 stoichiometry per GroEL toroid. This allowed both the position and the shape of dPJ9 in the GroEL/dPJ9 complexes to be determined. A single-ring GroEL/GroES variant complexed with one dPJ9 molecule was used to study the structural changes of dPJ9 in GroEL/GroES/dPJ9 complexes formed with ADP and with ATP. It was found that both the shape and the position of the bound dPJ9 in the GroEL/GroES/dPJ9 complex with ADP were the same as those in the GroEL/dPJ9 complex. However, dPJ9 assumed a more symmetric shape when bound in the GroEL/GroES/dPJ9 complex with ATP. This important observation reflects the relative ability of ATP to promote refolding of protein substrates relative to that of ADP.
Krueger
Susan
S
NIST Center for Neutron Research, National Institute of Standards and Technology, 100 Bureau Drive, Stop 8562, Bldg. 235/Room E151, Gaithersburg, MD 20899-8562, USA. susan.krueger@nist.gov
Gregurick
Susan K
SK
Zondlo
James
J
Eisenstein
Edward
E
D000244:Adenosine Diphosphate / Q000737:chemistry; D000255:Adenosine Triphosphate / Q000737:chemistry; D001412:Bacillus subtilis / Q000378:metabolism; D018835:Chaperonin 10 / Q000737:chemistry / Q000378:metabolism*; D018834:Chaperonin 60 / Q000737:chemistry / Q000378:metabolism*; D018833:Chaperonins / Q000737:chemistry; D004926:Escherichia coli / Q000378:metabolism; D017353:Gene Deletion; D008958:Models, Molecular; D009154:Mutation; D009502:Neutrons; D010957:Plasmids / Q000378:metabolism; D011485:Protein Binding; D017510:Protein Folding; D017434:Protein Structure, Tertiary; D012542:Scattering, Radiation; D020860:Subtilisin / Q000737:chemistry
D018835:Chaperonin 10; D018834:Chaperonin 60; D000244:Adenosine Diphosphate; D000255:Adenosine Triphosphate; D020860:Subtilisin; D018833:Chaperonins