Structural basis for the hyperthermostability of an archaeal enzyme induced by succinimide formation

Dongre, AV and Das, S and Bellur, A and Kumar, S and Chandrashekarmath, A and Karmakar, T and Balaram, P and Balasubramanian, S and Balaram, H (2021) Structural basis for the hyperthermostability of an archaeal enzyme induced by succinimide formation. In: Biophysical Journal .

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Abstract

Stability of proteins from hyperthermophiles (organisms existing under boiling water conditions) enabled by a reduction of conformational flexibility is realized through various mechanisms. A succinimide (SNN) arising from the post-translational cyclization of the side chains of aspartyl/asparaginyl residues with the backbone amide -NH of the succeeding residue would restrain the torsion angle Ψ and can serve as a new route for hyperthermostability. However, such a succinimide is typically prone to hydrolysis, transforming to either an aspartyl or β-isoaspartyl residue. Here, we present the crystal structure of Methanocaldococcus jannaschii glutamine amidotransferase and, using enhanced sampling molecular dynamics simulations, address the mechanism of its increased thermostability, up to 100°C, imparted by an unexpectedly stable succinimidyl residue at position 109. The stability of SNN109 to hydrolysis is seen to arise from its electrostatic shielding by the side-chain carboxylate group of its succeeding residue Asp110, as well as through n � �� interactions between SNN109 and its preceding residue Glu108, both of which prevent water access to SNN. The stable succinimidyl residue induces the formation of an α-turn structure involving 13-atom hydrogen bonding, which locks the local conformation, reducing protein flexibility. The destabilization of the protein upon replacement of SNN with a Φ-restricted prolyl residue highlights the specificity of the succinimidyl residue in imparting hyperthermostability to the enzyme. The conservation of the succinimide-forming tripeptide sequence (E(N/D)(E/D)) in several archaeal GATases strongly suggests an adaptation of this otherwise detrimental post-translational modification as a harbinger of thermostability. © 2021 Biophysical Society

Item Type:	Journal Article
Publication:	Biophysical Journal
Publisher:	Biophysical Society
Additional Information:	The copyright for this article belongs to Biophysical Society
Department/Centre:	Division of Biological Sciences > Molecular Biophysics Unit
Date Deposited:	21 Nov 2021 16:28
Last Modified:	21 Nov 2021 16:28
URI:	http://eprints.iisc.ac.in/id/eprint/69936

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