ProPrep: An Interactive and Instructional Interface for Proper Protein Preparation with AMBER.

Millions of experimental and AI-predicted protein structures are now available, and the biosynthetic promise of bespoke proteins is increasingly within reach. The functional characterization challenge thus posed cannot be addressed by experimental techniques alone. Molecular dynamics (MD) simulations offer functional screening with atomic resolution, yet accessibility remains limited. Existing computational chemistry software presents stark trade-offs whereby powerful tools require extensive expertise and manual effort, or user-friendly programs function as black boxes that obscure critical preparation decisions. Herein, we present ProPrep, an interactive workflow manager that guides users through expert-quality MD preparation by showing the 'what, why, and how' of each step while automating tedious manual operations. Within a single workspace, ProPrep integrates (1) downloading structures from multiple sources (PDB, AlphaFold, AlphaFill), (2) performing homology searches, (3) aligning structures, (4) curating and repairing structural issues, (5) applying mutations, (6) parameterizing specialized residues, (7) converting redox-active sites to forcefield-compatible forms, (8) generating topology and coordinate files, and (9) configuring, executing, and analyzing simulations with active monitoring of key quantities via ASCII visualizations. A key innovation is ProPrep's extensible transformer framework for detecting, defining, and transforming redox-active sites-including mono- and polynuclear metal centers, organic cofactors, and redox-active amino acids-for forcefield compatibility. We demonstrate the full workflow on a 64-heme cytochrome 'nanowire' bundle (PDB: 9YUQ), proceeding from a PDF file to energy minimization of the solvated system (467,635 atoms) for constant pH molecular dynamics-a process demanding 4,819 PDB record modifications and 610 bond definitions-in 18 minutes of user interaction. The entire process is recorded in an interactive session log that can be shared and replayed for reproducibility, making simulation setup a fully transparent process that relies on what was done instead of what was remembered and reported.