Genetic analysis of a weak D100 type individual produced anti-D
Objective:To perform genetic analysis of a RHD variant,predict and analyze the effect of the mu-tation on RhD antigen and evaluate the clinical significance of the variant type.Methods:Ten exons and adjacent flanking intron regions of RHD gene were amplified and analyzed by direct sequencing.RHD zygosity test was performed using Polymerase Chain Reaction-Sequence Specific Primers(PCR-SSP).PyMOL software were used to analyze 3D structures of RhD caused by the variant alleles.The effect of non-synonymous substitution was pre-dicted using Protein Variation Effect Analyzer(PROVEAN),Sorting Intolerant From Tolerant(SIFT)and Poly-morphism Phenotyping algorithm(PolyPhen-2)software.Results:The variant was hemizygous RHD(-).One single-nucleotide missense variant c.787G＞A was detceted in exon 5 encoding a p.G263R substitution.PyMOL 3D structural simulation analysis showed that hydrogen bonds between arginine and adjacent amino acids became shorter and increased compared with wild type.Two of the three bioinformatics programs suggested that p.G263R substitution was deleterious to RhD protein function.Conclusion:The RHD variant was classified as weak D100 type.The amino acid substitution caused by c.787G＞A mutation may affect the normal assembly of tertiary structure,leading to changes in RhD antigen characteristics.These results suggest that the D variant has the po-tential to produce anti-D when immunized with normal D antigen.
Rh blood groupD-variantweak D type 100PyMOLbioinformatics analysis
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NSTL
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