Abstract
Inactive mammalian tolloid-like 1 (tll1) and mutations detected in tolloid-like 1 (TLL1) have been linked to the lack of the heart septa formation in mice and to a similar human inborn condition called atrial-septal defect 6 (ASD6; OMIM 613087, formerly ASD II). Previously, we reported four point mutations in TLL1 found in approximately 20% of ASD6 patients. Three mutations in the coding sequence were M182L, V238A, and I629V. In this work, we present the effects of these mutations on TLL1 function. Three recombinant cDNA constructs carrying the mutations and one wild-type construct were prepared and then expressed in HT-1080 cells. Corresponding recombinant proteins were analyzed for their metalloendopeptidase activity using a native substrate, chordin. The results of these assays demonstrated that in comparison with the native TLL1, mutants cleaved chordin and procollagen I at significantly lower rates. CD analyses revealed significant structural differences between the higher order structure of wild-type and mutant variants. Moreover, biosensor-based assays of binding interactions between TLL1 variants and chordin demonstrated a significant decrease in the binding affinities of the mutated variants. The results from this work indicate that mutations detected in TLL1 of ASD6 patients altered its metalloendopeptidase activity, structure, and substrate-binding properties, thereby suggesting a possible pathomechanism of ASD6.
Highlights
Mammalian tolloid-like 1 (TLL1) is classified in the evolutionarily conserved family of zinc-dependent metalloendopeptidases
The first discovered tolloid of the family in mammals was bone morphogenetic protein 1 (BMP1) [2], known as procollagen C-endopeptidase, which engages in the formation of the extracellular matrix (ECM) [3,4]
The murine Tll−/− heart abnormalities reported by others [19] mimic those found in atrial-septal defect 6 (ASD6) in human, which is an inborn defect
Summary
Mammalian tolloid-like 1 (TLL1) is classified in the evolutionarily conserved family of zinc-dependent metalloendopeptidases. These metalloendopeptidases are known as tolloids (BMP1/TLD/Tolloid-like). BMP1 and the longer alternatively spliced variant of the BMP1 gene product, TLD [5], demonstrate high homology to a protein named Drosophila tolloid-related (tlrd) in fruit flies. The Drosophila tlrd is involved in embryonal signaling and development, leading to the designation of the dorsal–ventral body axis [6] Another protein, tolloid-related protein 1 (TLR1), is a key factor in transformation of the fly larvae [7,8]. The two other proteins (mammalian tolloid-like 1 [TLL1] and mammalian tolloid-like 2 [TLL2]) are encoded by separate genes [16,17,18]
Sign-in/Register to view highlights & summary Sign-in/Register to access full text options 
Talk to us
Join us for a 30 min session where you can share your feedback and ask us any queries you have
Schedule a callDisclaimer: All third-party content on this website/platform is and will remain the property of their respective owners and is provided on "as is" basis without any warranties, express or implied. Use of third-party content does not indicate any affiliation, sponsorship with or endorsement by them. Any references to third-party content is to identify the corresponding services and shall be considered fair use under The CopyrightLaw.
