A novel RHD allele (c.727 T>G) associated with reduced D antigen expression in a pregnant Chinese woman

This study aimed to characterize a novel RHD variant and to predict its potential impact on RhD antigen expression by integrating serological testing, molecular genotyping and structural modelling.

The full-length RHD gene was analysed using PacBio single-molecule real-time (SMRT) long-read sequencing. The effect of the identified variant on protein stability was evaluated using DynaMut2. Amino acid interactions, including hydrogen bonds and hydrophobic contacts, were analysed with LigPlot to illustrate two-dimensional (2D) interaction changes, while three-dimensional (3D) visualization and structural interpretation were performed using PyMOL.

Serological testing and sequencing revealed that the proband was heterozygous, with the reported RHD*15 allele and a novel allele carrying the c.727T>G (p.Tyr243Asp) variant present in trans. Structural prediction suggested that the p.Tyr243Asp substitution weakened hydrophobic interactions and reduced protein stability.

The coexistence of the reported RHD*15 allele and a novel missense variant, c.727T>G (p.Tyr243Asp), on different alleles may jointly contribute to reduced RhD antigen expression by affecting protein stability and epitope configuration. Integrating molecular genotyping and structural modelling with conventional serology provides complementary insights into the characterization of novel RHD variants and may provide additional clues for refining the classification of weak D and partial D phenotypes.