Discrimination of three mutational events that result in a disruption of the R122 primary autolysis site of the human cationic trypsinogen (PRSS1) by denaturing high performance liquid chromatography

BMC Genet. 2001;2:19. doi: 10.1186/1471-2156-2-19. Epub 2001 Nov 19.

Abstract

Background: R122, the primary autolysis site of the human cationic trypsinogen (PRSS1), constitutes an important "self-destruct" or "fail-safe" defensive mechanism against premature trypsin activation within the pancreas. Disruption of this site by a missense mutation, R122H, was found to cause hereditary pancreatitis. In addition to a c.365G>A (CGC>CAC) single nucleotide substitution, a c.365 through 366GC>AT (CGC>CAT) gene conversion event in exon 3 of PRSS1 was also found to result in a R122H mutation. This imposes a serious concern on the genotyping of pancreatitis by a widely used polymerase chain reaction-restriction fragment length polymorphism assay, which could only detect the commonest c.365G>A variant.

Materials and methods: DNA samples containing either the known c.365G>A or c.365 through 366GC>AT variant in exon 3 of PRSS1 were used as positive controls to establish a denaturing high performance liquid chromatography (DHPLC) assay.

Results: DHPLC could readily discriminate the two known different mutational events resulting in the R122H mutation. More importantly, under the same experimental conditions, it identified a further mutational event that also occurs in the R122 primary autolysis site but results in a different amino acid substitution: c.364C>T (CGC>TGC; R122C).

Conclusions: A rapid, simple, and low-cost assay for detecting both the known and new mutations occuring in the R122 primary autolysis site of PRSS1 was established. In addition, the newly found R122C variant represents a likely pancreatitis-predisposing mutation.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Amino Acid Substitution
  • Chromatography, High Pressure Liquid / methods*
  • DNA / chemistry
  • DNA Mutational Analysis / methods*
  • Gene Conversion
  • Genetic Predisposition to Disease
  • Humans
  • Mutation, Missense
  • Nucleic Acid Denaturation
  • Pancreatitis / genetics*
  • Trypsin*
  • Trypsinogen / chemistry
  • Trypsinogen / genetics*
  • Trypsinogen / metabolism

Substances

  • Trypsinogen
  • DNA
  • PRSS1 protein, human
  • Trypsin

Associated data

  • OMIM/167800
  • OMIM/276000