TIGP (BIO)—Structure determination beyond the reach of AlphaFold: a case of an intrinsically disordered protein deteriorating amyloidogenesis-related neurodegenerative diseases

Abstract

We recently proposed a novel scheme for determining the structures of intrinsically disordered proteins (IDPs) with a systemic structure enumeration method, named Threading-Augmented Interval Branch-and Pruned (TAiBP), which is based on a distance geometry algorithm and can be used forge the information from nuclear magnetic resonance (NMR) and small angle X-ray scattering (SAXS) experiments, to achieve very comprehensive generation of protein structures.  For the first time, the structure determination of the human SERF1a protein has been carried out with this new approach. SERF1a belongs to the MOAG-4/SERF class of proteins, which is a positive regulator of the aggregate formation of amyloid proteins and has been suggested to play important roles in numerous aging-related neurodegenerative diseases. Previous studies indicated that SERF1a is a very flexible protein, containing at least one α-helical region. Here we had provided the first description of the conformational ensembles of SERF1a at two pH values (6 and 6.8). We also compared the conformation generations for the case of SERF1a using CYANA-FLYA procedure and the TAiBP procedure. In our integrated scheme the generated conformations were filtered using the Pepsi-SAXS program by fitting the intensive curves from the SAXS experiments combining with size exclusion chromatography (i.e., SEC-SAXS).  At pH 6.8, a good fit of the SEC-SAXS curves can be obtained with the CYANA and TAiBP conformations, while at pH 6, the NMR conformations from the CYANA procedure cannot match the SEC-SAXS curves with both Pepsi-SAXS and WAXSiS, implying a problem of limited conformation generation due to the additional long-range nuclear Overhauser effects (NOEs) between residues Lys¹³ and Thr³² in this pH condition. The conformations generated by the TAiBP scheme indicated that the shortening of the C-terminal α-helix, as well as the destabilization of the N-terminal α-helix at acidic pH, may be related to the physiological function of SERF1a in the nucleoli. Interestingly, the conformations generated by TAiBP demonstrated that the N-terminal region of SERF1a displays numerous possible binding pockets, particularly in the region interacting with α-synuclein, as assessed by chemical shift perturbations. Our recent NMR-based screening using F¹⁹ compounds also discovered several compounds that can bind significantly with SERF1a, supporting the existences of these binding pockets. The conformations of SERF1a generated by TAiBP can also be used to derive a description of the internal dynamics in agreement with NMR relaxation. In general, the TAiBP procedure allows for a more comprehensive exploration conformational space which enables consistent interpretation of multifaceted biophysical characterization.
 
 

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