{'fraction_non_alphanumeric': 0.03033131124591694, 'fraction_numerical': 0.0055996266915538965, 'mean_word_length': 5.1082621082621085, 'pattern_counts': {'":': 0, '<': 0, '<?xml version=': 0, '>': 3, 'https://': 0, 'lorem ipsum': 0, 'www.': 0, 'xml': 0}, 'pii_count': 0, 'substrings_counts': 0, 'word_list_counts': {'cursed_substrings.json': 0, 'profanity_word_list.json': 0, 'sexual_word_list.json': 0, 'zh_pornsignals.json': 0}}
Interactions of Indole Alkaloids with Myoglobin: A Mass Spectrometry-based Spectrometric and Computational Method.

RATIONALE
Interactions of drug molecules and proteins play an important role in physiological and pathological processes in vivo. It is of significance to establish a reliable strategy for studying protein-drug ligand interactions and would be helpful for the design and screening of new drugs in pharmacological research.


METHODS
The interactions between four indole alkaloids (IAs) extracted from Ophiorrhiza japonica and myoglobin (Mb) protein were investigated by using a multi-spectrometric and computational method of native electrospray ionization mass spectrometry (Native ESI-MS), hydrogen/deuterium exchange mass spectrometry (HDX-MS), circular dichroism (CD) and molecular docking (MD).


RESULTS
The IA bound-Mb complexes were analyzed by native ESI-MS, with the obtained stoichiometry of protein-ligand at 1:1, 1:2, 1:3, respectively. Binding constants were measured according to the interpretation of MS spectra. MD complemented MS measurement, probed that the binding sites and modes of four IAs to Mb. Analyses of CD and HDX-MS demonstrated that the exposure to IAs could affect the conformation of Mb by decreasing the α-helix content and make Mb more susceptible to HDX at the backbone.


CONCLUSIONS
A new mass spectrometry-based integrated analysis method has been developed to successfully study the interactions of Mb and IAs extracted from Ophiorrhiza japonica. The experimental and calculation results have good consistency, revealing all of the four IA molecules could bind to Mb to form 1:1, 1:2 and 1:3 Mb-IA complexes, respectively. The order of binding ability of these IAs to Mb was Ophiorrhine B > Compound C > Ophiorrhine A > Compound D. CD and HDX-MS results indicated that binding with IAs destabilizes resulted of Mb. HDX-MS analysis suggests that Mb becomes more accessible to HDX, indicating that IAs binding destabilizes the structure of Mb. In addition, interacting with IAs affected the overall structure of Mb, ascribed to the decrease of α-helix content and less folding of backbone.