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Molecular recognition of ubiquitin and Lysine 63 linked diubiquitin by STAM2 : the effect of the linkers length and flexibility

Abstract : Protein-protein interaction is considered as an important field of research, as it is the key to control variable cell processes and pathways. In eucaryotic cells, multidomain proteins (MDPs), which consist of more than one domain, take up over 70 % of the pool. Those identical or different domains of a MDP are connected to each other by a linker of variable length and flexibility. For long flexible linker, it allows the protein to sample a wide range of conformation and to adjust interaction in a subtle way. Despite numerous efforts of research on the field, some issues remain unanswered or require further investigation. As part of this thesis, my work aims to define the role taken by the intrinsically disordered linker within MDPs. For that purpose, the STAM2 (Signal transducing adapter molecule 2) protein of the ESCRT (Endosomal Sorting Complexes Required for Transport) machinery was chosen to examine the effect of the flexibility and dynamics of the linker regions on the molecular recognition with ubiquitin and Lysine63-linked di-ubiquitin (K63-Ub2). Such efforts were carried out by designing specific mutants altering the linker regions in different ways. The various truncated versions undergo half or complete deletion of a domain or have their linker either shortened, deleted or modified in the amino acid composition. With a combination of the several biophysical methods namely NMR (Nuclear Magnetic Resonance) spin relaxation, SAXS (Small Angle X-ray Scattering) and CD (Circular Dichroism), the study has demonstrated that the alteration in the linker region modifies the flexibility and the dynamics of the protein, one among them possibly introduces slight change in conformation. Furthermore, the modification of the linker has an impact on the inter-domain motion and alter binding affinities between STAM2 constructs and di-ubiquitin without affecting domains integrity or binding sites. In brief, disordered linkers provide plasticity to the protein, which allow adaptability and specificity to molecular recognition process. As a further application, the linkers included in multidomain proteins could also be the next generation of druggable target as their modification may reduce or completely abolish interactions
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  • HAL Id : tel-03091978, version 1

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Minh-Ha Nguyen Thi. Molecular recognition of ubiquitin and Lysine 63 linked diubiquitin by STAM2 : the effect of the linkers length and flexibility. Other. Université de Lyon, 2019. English. ⟨NNT : 2019LYSE1321⟩. ⟨tel-03091978⟩

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