Biochemistry of Factor IX and Molecular Biology of Hemophilia B, страница 10

Smaller deletions of the factor IX gene cause hemophilia B. Factor IX Chicago-1 possesses a complex deletion of exons 5, 7, and 8, totaling approximately 25 kb. MLID87138319  153 Factor IX Seattle-1 contains an intragenic deletion of exons 5 and 6 of the factor IX gene, MLID86086362  154 resulting in the production of a truncated factor IX protein of 36,000 MW that is excreted into the urine. MLID86168884  155 Factor IX Yemen MLID89300787  149 and factor IX Tubingen MLID89300787  149 each possess deletions of exons 1–3 of the factor IX gene, with variable amounts of the 5'-flanking sequence also having been deleted. Factor IX Hanover has a deletion of exons 4 and 5 totaling 8 kb. MLID89300787  149 Factor IX Strasbourg contains a 2.8-kb intragenic deletion encompassing exon 4, the exon encoding the first EGF-like domain. MLID87000976  156 Despite possessing 30% of normal factor IX antigen levels, this family displays a severe hemophilic phenotype. Factor IX Ratingen shows a loss of exon 7, with a deletion of 1.5 kb. MLID89300787  149 Factor IX London-10 contains a deletion of the codon for Arg 37, MLID89305505  62 resulting in a severe hemophilic phenotype despite the presence of 12% of normal factor IX antigen levels. Factor IX Seattle-2 contains a deletion of a single adenine nucleotide at position 17,699 (in exon 4) of the factor IX gene, corresponding to residue 85 of the factor IX molecule. MLID88008341  157 This results in a frameshift mutation, creating a stop codon at codon 86. Factor IX London-11 (deletion of bases 31,059–31,060), factor IX London-12 (deletion of nucleotide 6392), and factor IX Malmo-1 (deletion of nucleotides 30,950–30,957) also result in frameshifts leading to premature stop codons. MLID89305505  62 It is unclear whether these mRNAs are processed normally, resulting in a truncated plasma protein secreted in the urine as above, or whether these frameshift mutations result in an unstable mRNA, producing a thalassemia-like syndrome.

In other cases, point mutations can lead to the production of little to no factor IX antigen. Rees et al. 158 and Winship 159 have described point mutations within the obligatory donor splice junction of exons 6 and 3, respectively. These mutations result in the production of an improperly spliced mRNA, causing a defect in transcription that results in no factor IX antigen being produced by these patients. Factor IX Bordeaux (Lys 411 B Stop), MLID89233115  160 factor IX Portland (Arg 252 B Stop), 161 factor IX Malmo-3 (Arg 248 B Stop), MLID89305505  62 Malmo-4 (Arg 29 B Stop), MLID89305505  62 and Malmo-5 (Trp 194 B Stop), MLID89305505  62 factor IX New York (Arg 338 B Stop), MLID89323424  162 factor IX Oxford 11 (Gln 11 B Stop), 163 factor IX Bonn-1 (Arg 338 B Stop), MLID89300787  149 an unnamed mutant (Arg 252 B Stop), MLID89118146  164 and other mutations all possess nonsense mutations resulting in premature stop codons. In each case, the premature stop signal yields an antigen-negative phenotype.

Finally, several missense mutations have been described that result in markedly reduced levels of factor IX antigen (Fig. 107-9). Factor IX London-8 (Cys 336 B Arg) results in the loss of a cysteine that is conserved not only in the coagulation proteases but in the digestive serine proteases as well. MLID89305505  62 This loss disrupts the tertiary structure of the catalytic domain. Factor IX London-9 (Asn 120 B Tyr) also leads to a severe, antigen-negative phenotype. MLID89305505  62 This substitution of a bulky, hydrophobic aromatic ring for the hydrophilic asparagine residue is presumed to yield an unstable protein product. Factor IX Oxford-5 (Gly 114 B Ala), 163 Oxford-8 (Glu 7 B Asp), 163 and Oxford-9 (Ala 291 B Pro) 163 all possess little circulating factor IX antigen.

A gene insertion and a combination of a gene insertion plus gene deletion have been reported to cause hemophilia B. Hemophilia B El Salvador contains a 6.1-kb insertion near exon 4 of the factor IX gene, MLID88161065  165 resulting in moderate hemophilia, with factor IX activity of 1% and factor IX antigen of 6%. A combination of a 2-kb insertion and a 1-kb deletion mutation in intron 6 results in hemophilia B Sydney, in which there is no circulating factor IX antigen. 166

Antigen-Positive Hemophilia B