The CRIPTO/FRL‐1/CRYPTIC (CFC) domain of human Cripto

Abstract

The disulfide structure of the CRIPTO/FRL-1/CRYPTIC (CFC) domain of human Cripto protein was determined by a combination of enzymatic and chemical fragmentation, followed by chromatographic separation of the fragments, and characterization by mass spectrometry and N-terminal sequencing. These studies showed that Cys115 forms a disulfide bond with Cys133, Cys128 with Cys149, and Cys131 with Cys140. Protein database searching and molecular modeling revealed that the pattern of disulfide linkages in the CFC domain of Cripto is the same as that in PARS intercerebralis major Peptide C (PMP-C), a serine protease inhibitor, and that the EGF-CFC domains of Cripto are predicted to be structurally homologous to the EGF-VWFC domains of the C-terminal extracellular portions of Jagged 1 and Jagged 2. Biochemical studies of the interactions of ALK4 with the CFC domain of Cripto by fluorescence-activated cell sorter analysis indicate that the CFC domain binds to ALK4 independent of the EGF domain. A molecular model of the CFC domain of Cripto was constructed based on the nuclear magnetic resonance structure of PMP-C. This model reveals a hydrophobic patch in the domain opposite to the presumed ALK4 binding site. This hydrophobic patch may be functionally important for the formation of intra or intermolecular complexes.

Solution Structure and Dynamics of a Prototypical Chordin-like Cysteine-rich Repeat (von Willebrand Factor Type C Module) from Collagen IIA

“…Other disulfide bond patterns were excluded based on the absence of NOEs. These experimental data contradict the predicted disulfide bond pattern for the CR repeat, based on sequence comparison with the CFC domain from Cripto (66).…”

A Novel Database of Disulfide Patterns and its Application to the Discovery of Distantly Related Homologs

“…28 Cripto is a protein involved in early embryonic development and was shown to be overexpressed in a number of human cancers. 29 Cripto family proteins are characterized by two cysteine-rich structural motifs: an epidermal growth factor (EGF)-like domain and a CFC domain, the latter of which is considered unique to this family.…”

Current View on EpCAM Structural Biology

“…Likewise, the spacing between cysteine residues alongside the polypeptide chain differs between EGF-like domains and EpCAM ND. However, the ND does share disulfide linkage with some other small cysteine-rich domains, like the CFC domain of human cripto [ 18 ], and is with regard to fold [ 19 ] most similar to the disulfide-free WW-type domains [ 20 ] found in various proteins like formin-binding protein 3, dystrophin and NEDD, still, all these combined structural aspects are not found in any other protein of known structure, thereby classifying the structure of EpCAM N -terminal domain as unique. Domains of this type, also called cysteine-rich microdomains (size up to 40 aa residues), are often involved in protein–protein interactions, an example is a well-known EGF–EGFR interaction [ 21 ], and are also considered as therapeutic scaffolds with a very stable central core [ 22 ].…”