Elucidating the link between NS-SNP-Induced protein conformational changes and drug resistance in Klebsiella pneumoniae.

The escalating drug-resistance of Klebsiella pneumoniae (K. pneumoniae), a leading cause of both community-acquired and nosocomial infections, poses a severe threat to global public health. While non-synonymous single nucleotide polymorphisms (NS-SNPs) serve as crucial molecular markers in microbial resistance studies, their role in inducing protein conformational changes to confer resistance in K. pneumoniae remains unexplored. This study integrates machine learning with protein structural analysis to elucidate the link between NS-SNP-mediated conformational alterations in homologous proteins and multidrug resistance in K. pneumoniae. This work aligned whole-genome sequences against the K. pneumoniae HS11286 reference genome using MUMmer 3 to generate SNP datasets. Two custom machine learning algorithms, Fast Feature Selection (FFS) and Codon Mutation Detection (CMD), were employed for SNP feature extraction and NS-SNP identification, respectively. Subsequently, ab initio method and homology modeling were conducted on NS-SNP-modified amino acid sequences, followed by protein quality assessment. Then, a statistical association analysis of genotype-phenotype was performed. Structural comparisons were performed using Molecular Operating Environment (MOE) software, and cross-validation of protein modeling was performed using AlphaFold2. Finally, the electrostatic surface potential alterations were analyzed via PyMOL. The results identify four NS-SNP mutations (IDs 1208814, 3086795, 3509283, and 3662328) whose mediated protein conformational changes are strongly associated with resistance. Notably, this study found that the porin variation (porin loss, porin channel narrows, and low expression of porin) were closely related to the NS-SNP (ID 3662328) mutation. Electrostatic potential analysis suggests that resistance may stem from mutation-induced changes in electrostatic energy, implying that an altered electrostatic environment could hinder antibiotic binding affinity. This elucidates a bacterial resistance mechanism related to NS-SNP mutations, and provides a valuable foundation for clinical research and therapeutic strategies against K. pneumoniae infections.