The epidermal growth factor precursor (proEGF) is a large membrane protein containing nine EGF units in the extracellular domain. The EGF unit closest to the membrane corresponds to mature EGF. Proteolytic cleavage at the C-terminus of the extracellular domain generates a soluble fragment. Several other membrane-anchored growth factors, members of the EGF gene family, undergo similar proteolytic cleavage and subsequent release. The proteases responsible for this growth factor maturation have not yet been identified. Some information is available regarding the amino acid sequence bounding the putative cleavage site. A consensus oligonucleotide primer encoding this region and a primer encoding part of the EGF unit has been used for PCR screening of several lambda gt11 cDNA libraries as a first step to determine the ubiquity of this cleavage site. The site appears to be shared by several membrane-anchored proteins encoded in a salivary gland cDNA library. Construction of more specific primers is underway. Additionally, construction of a rat salivary gland cDNA library in lambda ZAP will afford use of the pBluescript phagemid. Once constructed, this library will be screened by PCR for both the putative cleavage site and any known protein convertases. Other studies using PCR have identified several inducible protein convertases in two other libraries, one from heart and one from pituitary. These convertases may be involved in the proteolytic cleavage. Further work needs to be done.
