A DOB allele encoding an amino acid substitution (Phe62Ser) resulting in a Dombrock null phenotype.
BACKGROUND
The gene polymorphisms responsible for the antigens Doa, Dob, Hy, and Joa in the Dombrock (Do) blood group system have been identified. Four different mutations have been reported to cause the Dombrock null [Gy(a-)] phenotype. These include splice mutations, an eight-nucleotide deletion, and insertion of a stop codon. Here a Dombrock null caused by a single-amino-acid substitution in the full-length protein is reported.


METHODS
DOA and DOB were determined by polymerase chain reaction-restriction fragment length polymorphism, and DO (ART4) exons and flanking regions were sequenced from genomic DNA. Expression analysis was performed by transfection of wild-type and mutant cDNAs into HEK 293T cells followed by flow cytometry and immunoblotting. Homology modeling was used to map the mutation on the protein structure.


RESULTS
The patient's sample carried nt 793G/G, indicating a DOB/DOB background. Exon 2 sequencing showed the sample carried a new mutation, nt 185T>C, causing a Phe62Ser substitution. This variant Do was not expressed on the surface of transfected HEK 293T cells. The mutation maps to a highly conserved FDDQY motif located between the beta1-strand and alpha1-helix near the COOH terminus in the native molecule.


CONCLUSIONS
The Dombrock null reported here is due to a single Phe62Ser mutation. The expression data confirmed that 62Ser is responsible for lack of cell surface Do, and protein modeling suggests the mutation disrupts important aromatic side chain interactions between Phe62 and His160. Production of an antibody to a high prevalence Dombrock antigen (anti-Gya) in this patient was consistent with complete absence of Dombrock/ART4 protein.
Westhoff
Connie
C
American Red Cross, Philadelphia, PA 19123, USA. westhoffc@usa.redcross
Vege
Sunitha
S
Yazdanbakhsh
Karina
K
Wylie
Dwane
D
Razib
Mohammad
M
Hue-Roye
Kim
K
Halverson
Gregory
G
Read
Sandy
S
Whiteoak
Elizabeth
E
Nickle
Pam
P
Maurer
Joan
J
Kavitsky
Donna
D
Nance
Sandra
S
Reid
Marion E
ME
D036002:ADP Ribose Transferases / Q000737:chemistry / Q000235:genetics*; D000368:Aged; D000483:Alleles; D020816:Amino Acid Motifs; D019943:Amino Acid Substitution; D002460:Cell Line; D005260:Female; D006801:Humans; D008565:Membrane Proteins / Q000737:chemistry / Q000235:genetics*; D008958:Models, Molecular; D010641:Phenotype; D016133:Polymerase Chain Reaction; D012150:Polymorphism, Restriction Fragment Length; D017422:Sequence Analysis, DNA
D008565:Membrane Proteins; D036002:ADP Ribose Transferases; C105234:ART4 protein, human