Glycoconjugate Journal (1996) 13:985-993 Molecular basis for erythrocyte Le(a+b+) and salivary ABH partial-secretor phenotypes: expression of a FUT2 secretor allele with an A-+T mutation at nucleotide 385 correlates with reduced tt(1,2)fucosyltransferase activity STEPHEN HENRY 1. 1' ROSELLA MOLLICONE 2, PILAR FERNANDEZ 2 BO SAMUELSSON , RAFAEL ORIOL 2 and GORAN LARSON 1 IDepartment of Clinical Chemistry and Transfusion Medicine, Gdteborg University, Sweden 2INSERM U-178, UniversiO~ of Paris South (AT), Villejuif 94807, France Received 19 February 1996, revised 3 April 1996, accepted 6 April 1996 The Se ~,As85T mutation of the FUT2 gene was found to correlate with both the erthrocyte Le(a+b+) and/or salivary ABH partial-secretor phenotypes of Polynesians. Constructs with FUT1 and FUT2 wild type genes, and the FUT2 Sewa385T, seQ428A and sec571T mutated alleles, were cloned into pcDNAI, and expressed in COS-7 cells. COS-7 cells transfected with the Se ~vA385T allele had weak, but detectable, ct(1,2)fucosyltransferase activity, with an acceptor substrate pattern similar to the wild type FUT2 gene. Comparative kinetic studies from cell extracts with mutated SewA3sSTand wild type FUT2 alleles gave similar Km values, but less enzyme activity was present in cells transfected with Se wA385T (//max 230pmolh 1mg-1), as compared to those transfected with FUT2 .(Vma• 1030 pmolh -1 mg-1), suggesting that the mutated enzyme is more unstable. These results confirm that the molecular basis for the erytba'ocyte Le(a+b+), and the associated ABH salivary partial-secretor phenotype, is an amino acid change of Ile129 -+ Phe in the secretor a(1,2)fucosyltransferase. Keywords: secretor, fucosyltransferase, Lewis a, Lewis b, saliva ABH, blood group, FUT2, ABO, Polynesia Abbreviations: a(1,3/1,4)fucosyltransferase, GDP-L-fucose:fi-D-N-acetylglucosaminide 3/4-ct-L-fucosyltransferase; a(1,2)Ncosyltransferase, GDP-L-fucose: fi-D-galactoside-2-a-L-fucosyltransferase; bp, base pairs; FUT1, H gene; FUT2, Se gene; FUT3, Lewis gene or Fuc-TIII gene; FUT4, Fuc-TIV gene; FUT5, Fuc-TV gene; FUT6, Fuc-TVI gene; MAb, monoclonal antibody; PCR, polymerase chain reaction; RFLR restriction fragment length polymorphism; seG428A, FUT2 nonsecretor G--+ A mutation at nucleotide 428; se C571T, FUT2 nonsecretor C--+ T mutation at nucleotide 571; SeWA385r, FUT2 secretor weak A--,T mutation at nucleotide 385; SSP, sequence specific primer. Introduction The Lewis histo-blood group system is constituted of two major serologically defined antigens, Le a and Le b (as reviewed in [1]). The two fucosyltransferase genes, FUT2 and FUT3, responsible for the synthesis of Le a and Le b glycoconjugates have been cloned, sequenced and ex- pressed [2, 3]. *To whom correspondence and reprint requests should be adchessed. 0282-0080 9 1996 Chapman & Hall As a consequence of polymorphism of the genetically independent Lewis and secretor systems and interaction of their transferases, four major red cell and salivary ABH secretor phenotypes are common in Europeans. These are: Lewis-negative secretors and nonsecretors, both red cell phenotyping as Le(a-b-); Lewis-positive nonsecretors whose red cell phenotype will be Le(a+b-); and Lewis-positive secretors whose red cell phenotype will be Le(a-b+). There is also another red cell phenotype known as Le(a+b+). This red cell phenotype