SEARCH FOR MUTATIONS IN A SEGMENT OF THE EXON-28 OF THE HUMAN VON-WILLEBRAND-FACTOR GENE - NEW MUTATIONS, R1315C AND R1341W, ASSOCIATED WITH TYPE 2M AND 2B VARIANTS

von Willebrand Disease (vWD) is the most frequently inherited bleeding disorder in humans, and is caused by a qualitative and/or quantitativ e abnormality of the von Willebrand factor (vWF), A large number of de fects that cause qualitative variants have been located in the Al doma in of the vWF, which contains sites for interaction with platelet glyc oprotein Ib (GPIb). We have developed a new approach to detect mutatio ns based on Ddel digestion and single-strand conformation polymorphism analysis. A segment of 487 nucleotides, extending from intron 27 to c odon 1368 of the pre-pro vWF was amplified from genomic DNA, The cleav age with Ddel yields two fragments of appropriate size for this kind o f analysis and confirms that the gene, rather than the pseudogene, is being investigated, Six families with type 2B vWD: one type 2M vWD fam ily, and one another type 2A vWD family were studied. After sequencing the fragments with an altered electrophoretic pattern, we found four mutations previously described-R1308C, V1316M, P1337L, and R1306W-in p atients with 2B vWD, The last one arose de novo in the patient. In add ition, two new candidate mutations were observed: R1315C and R1341W. T he first one was associated to type 2M vWD, whereas the one second cos egregated with type 2B vWD. The fact that these new mutations were not found in 100 normal alleles screened further supports their causal re lationship with the disease, These mutations, which induce either a ga in or a loss of function, further show an important regulatory role of this region in the binding of vWF to GPIb and its implications in cau sing disease. (C) 1998 Wiley-Liss, Inc.