Mammalian expression of full-length bovine aggrecan and link protein: Formation of recombinant proteoglycan aggregates and analysis of proteolytic cleavage by ADAMTS-4 and MMP-13

Effects of covalently attached chondroitin sulfate on aggrecan cleavage by ADAMTS-4 and MMP-13

“…We have also analyzed cleavage of CS-substituted and CS-deficient FLAG-rbAgg by MMP-13, which has no apparent GAG-binding motifs. We have previously shown that MMP-13 cleaves aggrecan preferentially within the CS-rich domain (Miwa et al, 2006). The present work demonstrated that CS covalently attached to the aggrecan core protein enhances cleavage within the CS-2 domain by ADAMTS-4 (p68).…”

“…The p40 form of ADAMTS-4, however, has essentially negligible cleavage activity within the CS-2 domain (Miwa et al, 2006). This observation led us to speculate that the mechanisms of cleavage within the IGD and the CS-2 domain are quite distinct.…”

“…We used enzymatically deglycosylated steer aggrecan isolated from cartilage as well as deglycosylated or biosynthetically modified FLAG-tagged recombinant bovine aggrecan (FLAG-rbAgg) (Miwa et al, 2006) as substrates for the p68 and the p40 isoforms of ADAMTS-4. FLAG-rbAgg substituted with or without CS was generated by transient transfection and expression in wild-type and xylosyltransferase-deficient CHO mutant cell lines.…”

Proteoglycans of reactive rat cortical astrocyte cultures: Abundance of N-unsubstituted glucosamine-enriched heparan sulfate

“…R040001). Bovine A1A1D1 aggrecan was prepared as previously described (Miwa et al, 2006). Sephadex G-50 (Cat.…”

Tissue Engineering by Molecular Disassembly and Reassembly: Biomimetic Retention of Mechanically Functional Aggrecan in Hydrogel

“…1 Since AGC monomers extracted from mature tissue maintain the ability to form aggregates in vitro 33 and PG aggregation is a highly conserved molecular mechanism across species, the approach described here is likely to be applicable to AGC from other sources, including human sources, and to other HA-binding PGs to create a biomimetic matrix that can be used as a homograft. Recombinant forms of AGC and LP, which have been produced and demonstrated to maintain the ability to form aggregates, 34,35 along with commercially available HA, also may provide other sources of purified PG components.…”