NMR chemical shift assignment of a constitutively active fragment of the antitermination protein LicT

Yinshan Yang , Nathalie Declerck (1) , Héléne Démèné (1)

1 CBS - Centre de Biochimie Structurale [Montpellier]

LicT belongs to an essential family of bacterial antitermination proteins which bind to nascent mRNAs in order to stimulate transcription of sugar-metabolizing operons. As most of other antitermination proteins involved in carbohydrate metabolism, LicT is composed of a N-terminal RNA-binding module (CAT) and two homologous regulatory modules (PRD1 and PRD2). The activity of the CAT effector module is controlled by antagonist phosphorylations by the phosphotransferase system on conserved histidines of the two C-terminal PRDs in response to available carbon sources. Previous studies on truncated and mutant constructs have provided partial structural insight into the mechanism of signal transduction between the N-terminal RNA-binding domain and the two regulation modules. However, no structure at atomic resolution has been ever solved that contain the RNA-binding domain and a regulation module. We report the NMR assignment of a constitutively active fragment of LicT, named D99A-CAT-PRD1 or CAT-PRD1*. This fragment is composed of the RNA-binding module and the first N-terminal regulation module which bears the mutation of Asp99 to an asparagine. It is dimeric as the native protein, with a 40 kD molecular weight. The D99N mutation is sufficient to endow this fragment with a high RNA-binding constitutive activity, in a phosphorylation-free context. The assignment reported here should set the base of future NMR investigation of signal transduction between the regulatory module and the effector module in the active state of the protein, and in the long term enable the structural study of the full length protein structure in interaction with its target RNA.

Domaines: Sciences du Vivant [q-bio]

Yinshan Yang, Nathalie Declerck, Héléne Démèné. NMR chemical shift assignment of a constitutively active fragment of the antitermination protein LicT. Biomolecular NMR Assignments, 2020, 14, pp.19-22. ⟨10.1007/s12104-019-09914-w⟩. ⟨hal-02322846⟩