(P-IG-34) Novel FUT1 variant c.789C>A in patient with para-Bombay phenotype and evaluation by protein modeling

Background/Case Studies: ABO antigen expression is dependent on three genes: FUT1 and FUT2 encode fucosyltransferase responsible for expression of H antigen on RBCs and in secretions, respectively; ABO gene encodes glycosyltransferase responsible for conversion of H to A, B antigens. The rare para-Bombay phenotype (weak or no A, B, H antigens on RBCs) results from FUT1 mutations. We investigated the ABO and FUT genes in a Hispanic patient whose RBCs were non-reactive to weakly reactive with anti-A, moderately reactive with anti-A,B and non-reactive with anti-A1 lectin and -H, and whose plasma showed anti-H reactivity. We used open access protein modeling software to assess the impact of the novel missense variant and previously reported similar variants on protein structure

Study

Design/Methods: DNA was isolated from white blood cells. Whole genome sequencing (WGS) was performed, and results analyzed with automated blood group antigen interpretive software bloodTyper. Sanger sequencing was performed for confirmation of variants. The protein structures of human FUT1 and FUT1 variants were predicted with open access AlphaFold2 and visualized with software, PyMOL, with comparison to structure of the Arabidopsis thaliana FUT1 homolog.

Results/Findings: WGS found homozygous ABO*A1.01; two FUT2 synonymous heterozygous changes c.390C>T and c.513C>T, and a FUT1 homozygous novel variant c.789C>A (p.Asn263Lys), rs751831069, with frequency of 0.00006541 in Latino/Admixed American population (gnomAD v3.1.2); FUT1/2 changes were confirmed by Sanger sequencing. Structural evaluation of FUT1 p.Asn263Lys with a structural model of human FUT1, with insights from FUT1 homolog, showed that this change was located near the GDP binding pocket within the catalytic domain. A systematic evaluation of previously reported FUT1 null missense changes found p.Asn263His (FUT1*01N.34) and p.Ser262Lys (FUT1*01N.11), also located near the GDP binding pocket.

Conclusions: We report a novel FUT1 variant, c.789C>A (p.Asn263Lys) and a normal FUT2 gene in a Hispanic patient with weak A, H- phenotype and anti-H reactivity in the plasma (ie. para-Bombay). Computational analysis of p.263Lys and null variants p.263His and p.262Lys places these residues in the catalytic domain. The changes from polar Asn and Ser to positively-charged Lys and His likely disrupt catalytic activity, explaining the H- phenotype. The observed weak A is due to adsorption of secreted A substance onto RBCs. The publicly available AlphaFold2 software can be of great interest to transfusion professionals to predict structural changes in blood group antigens and to evaluate potential correlations between such changes and immunogenicity.

Importance of research: ABO antigen expression is dependent on three genes: FUT1, FUT2, and ABO. Mutations to the FUT1 gene results in the rare para-Bombay phenotype (weak or no A, B, H antigens on RBCs). We investigated a sample referred with weak A, H-, and anti-H in the plasma and identified a novel homozygous FUT1 variant. Additionally, we used the publicly available protein modeling software AlphaFold2 to assess the impact of this novel variant and reported similar variants on protein structure.