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Age-dependent Structural Reorganization of the Human Plasma Proteome

Aging research has largely focused on changes in protein expression levels, but aging cannot be explained by abundance changes alone. Here we focus on an under-explored dimension: structural changes in proteins and the dynamics of their interaction networks. In this study, we integrated experimental proteomics with computational structural biology to comprehensively characterize the age-dependent structural dynamics of plasma proteins — that is, how plasma proteins change structurally over a healthy life course.

1. Data Preprocessing

Data_preprocessing_for_PAR.R Computes the Peak Area Ratio (PAR) for each ion, using the 6 ions from the 1st, 2nd, and 3rd precursors of peptide-03 among the internal standards (IS), in order to select the IS used for normalization. (Fig. 1B — code added 2025-10-19)

PAR_distribution_histogram.R Applies a log10 transformation to the PAR1–6 values obtained above and plots their distribution as a histogram, together with the corresponding CV values. (Fig. 1A, C — code added 2025-10-19)

PAR_input_preprocessing.R

Aging_log_norm_data_preprocessing.ipynb

PAR_data_preprocessing_Jhyh.R Preprocesses the data required for the downstream analyses.

2. Statistical Analysis

Data_preprocessing_Jhyh+cosine similarity.R Performs cosine similarity analysis on the preprocessed data. (Fig. 2A, B)

limma_test_Jh_revised_added_fit3_final.R Uses limma to extract only the ions satisfying the fold-change and p-value criteria for each pattern, and plots the mean PAR change per pattern. (Fig. 3A, B)

overlap_pattern_fit3_final.R Merges the cosine similarity and limma results and retains only the overlapping ions.

overlap_boxplot_fit3_final.R Generates boxplots for each ion, grouped by pattern. (Fig. 3C)

3. Structural Analysis

pymol_interaction_distance.txt When inspecting AlphaFold3 multimer results in PyMOL, measures the minimum distance across the peptide– and protein–interface within each pattern's protein complex (1:1).

pymol_mean_plddt.txt When inspecting AlphaFold3 multimer results in PyMOL, computes the mean pLDDT. (Fig. 6B, C, D)

Notes

Cytoscape: Full STRING network, confidence ≥ 0.7, Homo sapiens (see manual for details). AF3 multimer web server: For PTMs, only N-linked glycosylation sites were added as NAG.

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Age-dependent Structural Reorganization of the Human Plasma Proteome

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