Mutations in the FKBP14 gene encoding FKBP22 (FK506 Binding Protein 22 kDa) cause kyphoscoliotic Ehlers-Danlos Syndrome (kEDS). The first clinical report showed that a lack of FKBP22 protein due to mutations causing nonsense-mediated decay of the mRNA leads to a wide spectrum of clinical phenotypes including progressive kyphoscoliosis, joint hypermobility, hypotonia, hyperelastic skin, hearing loss and aortic rupture. Our previous work showed that these phenotypic features could be correlated with the functions of FKBP22, which preferentially binds to type III, VI and X collagens, but not to type I, II or V collagens. We also showed that FKBP22 catalyzed the folding of type III collagen through its prolyl isomerase activity and acted as a molecular chaperone for type III collagen. Recently, a novel missense mutation Met48Lys in FKBP22 was identified in a patient with kEDS. In this report, we expand the list of substrates of FKBP22 and also demonstrate that the Met48Lys mutation diminishes the activities of FKBP22, indicating that pathology can arise from absence of FKBP22, or partial loss of its function.

Collagen is the most abundant protein in humans. It has a characteristic triple-helix structure and is heavily posttranslationally modified. The complex biosynthesis of collagen involves processing by many enzymes and chaperones in the rough endoplasmic reticulum. Lysyl hydroxylase 1 (LH1) is required to hydroxylate lysine for cross-linking and carbohydrate attachment within collagen triple helical sequences. Additionally, a recent study of prolyl 3-hydroxylase 3 (P3H3) demonstrated that this enzyme may be critical for LH1 activity; however, the details surrounding its involvement remain unclear. If P3H3 is an LH1 chaperone that is critical for LH1 activity, P3H3 and LH1 null mice should display a similar deficiency in lysyl hydroxylation. To test this hypothesis, we compared the amount and location of hydroxylysine in the triple helical domains of type V and I collagen from P3H3 null, LH1 null, and wild-type mice. The amount of hydroxylysine in type V collagen was reduced in P3H3 null mice, but surprisingly type V collagen from LH1 null mice contained as much hydroxylysine as type V collagen from wild-type mice. In type I collagen, our results indicate that LH1 plays a global enzymatic role in lysyl hydroxylation. P3H3 is also involved in lysyl hydroxylation, particularly at cross-link formation sites, but is not required for all lysyl hydroxylation sites. In summary, our study suggests that LH1 and P3H3 likely have two distinct mechanisms to recognize different collagen types and to distinguish cross-link formation sites from other sites in type I collagen.