Gene sequencing analysis and protein structural modeling for a case with Aw26 subtype of the ABO blood group.
10.3760/cma.j.cn511374-20240704-00372
- Author:
Qianqian CHEN
1
,
2
;
Jinrong CHEN
;
Kaizhao HUANG
;
Jiajin LIN
Author Information
1. The Second Affiliated Hospital and Yuying Children's Hospital of Wenzhou Medical University, Wenzhou, Zhejiang 325027, China. ljj88879099@
2. com.
- Publication Type:English Abstract
- MeSH:
ABO Blood-Group System/chemistry*;
Humans;
Molecular Dynamics Simulation;
Models, Molecular;
Mutation;
Genotype;
Protein Conformation;
Glycosyltransferases/chemistry*;
Male
- From:
Chinese Journal of Medical Genetics
2025;42(6):667-674
- CountryChina
- Language:Chinese
-
Abstract:
OBJECTIVE:To analyze the sequencing results, protein structure model, and impact of mutations on the dynamic stability of glycosyltransferase (GTA) in a case with Aw26 blood group subtype.
METHODS:ABO phenotype was determined by serological testing (anti-A, anti-B, anti-H, and reverse typing). Potential variant of the ABO gene was identified by Sanger sequencing, and the haploid sequence of the variant site was analyzed by TOPOT-A cloning. Molecular models of the GTA was generated by PyMol, and 100-ns molecular dynamics (MD) was simulated with GROMACS software to assess the conformational stability using root mean square deviation (RMSD), radius of gyration (Rg), solvent-accessible surface area (SASA), hydrogen bonding, and binding free energy.
RESULTS:Serological assays confirmed the proband as an Aw subtype, whose genotype was identified as ABO*Aw.26/ABO*O.01.02 with variants including p.Pro156Leu, p.Arg176His and p.Pro354ArgfsTer23. Haploid sequencing validated the results of direct sequencing. Molecular modeling showed that the p.Arg176His variant could reduce water-mediated hydrogen bonds from six (wild-type) to one (variant). MD simulation revealed the wild type system could achieve equilibrium within 10 ns (mean RMSD ≈ 0.30 nm), whilst the mutant system required 50 ns to equilibrate and exhibited greater fluctuation (mean RMSD ≈ 0.40 nm). Root mean square fluctuation (RMSF) analysis confirmed significantly increased flexibility in the mutant's N-terminal loop (residues 63-76). The mutant Rg displayed an expansion-contraction transition within 0 ~ 40 ns, and its SASA value has increased. The number of hydrogen bonds and binding energy of the mutant had decreased (wild-type: 5 to 8, binding energy: -11.53 kcal/mol; mutant: 2 to 5, binding energy:-8.52 kcal/mol).
CONCLUSION:An Aw26 subtype was identified. The p.Arg176His and p.Pro354Argfs*23p variants could synergistically compromise the structural stability of GTA and its substrate binding capacity by disrupting the hydrogen-bond network, increasing local flexibility, and reducing the overall conformational stability.
