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Dissociation of the Dimer of the Intrinsically Disordered Domain of RNase Y upon Antibody Binding.

Abstract：

:Although RNase Y acts as the key enzyme initiating messenger RNA decay in Bacillus subtilis and likely in many other Gram-positive bacteria, its three-dimensional structure remains unknown. An antibody belonging to the rare immunoglobulin G (IgG) 2b λx isotype was raised against a 12-residue conserved peptide from the N-terminal noncatalytic domain of B. subtilis RNase Y (BsRNaseY) that is predicted to be intrinsically disordered. Here, we show that this domain can be produced as a stand-alone protein called Nter-BsRNaseY that undergoes conformational changes between monomeric and dimeric forms. Circular dichroism and size exclusion chromatography coupled with multiangle light scattering or with small angle x-ray scattering indicate that the Nter-BsRNaseY dimer displays an elongated form and a high content of α-helices, in agreement with the existence of a central coiled-coil structure appended with flexible ends, and that the monomeric state of Nter-BsRNaseY is favored upon binding the fragment antigen binding (Fab) of the antibody. The dissociation constants of the IgG/BsRNaseY, IgG/Nter-BsRNaseY, and IgG/peptide complexes indicate that the affinity of the IgG for Nter-BsRNaseY is in the nM range and suggest that the peptide is less accessible in BsRNaseY than in Nter-BsRNaseY. The crystal structure of the Fab in complex with the peptide antigen shows that the peptide adopts an elongated U-shaped conformation in which the unique hydrophobic residue of the peptide, Leu6, is completely buried. The peptide/Fab complex may mimic the interaction of a microdomain of the N-terminal domain of BsRNaseY with one of its cellular partners within the degradosome complex. Altogether, our results suggest that BsRNaseY may become accessible for protein interaction upon dissociation of its N-terminal domain into the monomeric form.

journal_name

Biophys J

journal_title

Biophysical journal

authors

Hardouin P,Velours C,Bou-Nader C,Assrir N,Laalami S,Putzer H,Durand D,Golinelli-Pimpaneau B

doi

10.1016/j.bpj.2018.10.016

subject

Has Abstract

pub_date

2018-12-04 00:00:00

pages

2102-2113

issue

eissn

0006-3495

issn

1542-0086

pii

S0006-3495(18)31164-0

journal_volume

115

pub_type

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Dissociation of the Dimer of the Intrinsically Disordered Domain of RNase Y upon Antibody Binding.