The type XIII collagen ectodomain is a 150-nm rod and capable of binding to fibronectin, nidogen-2, perlecan, and heparin.
Type XIII collagen consists of a short N-terminal intracellular domain, a transmembrane domain, and a collagenous ectodomain, and it is found at many sites of cell adhesion. We report on the characterization of recombinant type XIII collagen. The shed ectodomain was purified from insect cell culture medium and shown to form 240-kDa trimers with a T(m) of 42 degrees C. Correct chain association into a triple-helical conformation was confirmed by limited pepsin digestion and CD spectroscopy. Rotary shadowing electron microscopy of the ectodomain revealed it to be a 150-nm rod with two flexible hinges separating 31-, 52-, and 68-nm portions. The rods represent the collagenous domains 1-3, and the hinges coincide with the non-collagenous domains 2 and 3. By using surface plasmon resonance analysis, the ectodomain showed interaction with immobilized fibronectin, nidogen-2, and perlecan with K(D) values in the nanomolar range. The binding sites of type XIII collagen for fibronectin were localized to the collagenous domains, whereas the binding activities for nidogen-2 and perlecan resided in the pepsin-sensitive portions of the ectodomain. Furthermore, the ectodomain bound significantly to heparin, which also inhibited shedding of the ectodomain in insect cell cultures. The results reveal that type XIII collagen is notably distinct in its structure compared with other cell-surface proteins, and the in vitro binding with fibronectin, heparin, and two basement membrane components is indicative of multiple cell-matrix interactions in which this ubiquitously expressed protein participates.
Tu
Hongmin
H
Collagen Research Unit, Biocenter and Department of Medical Biochemistry and Molecular Biology, University of Oulu, Aapistie 7, FIN-90220 Oulu, Finland.
Sasaki
Takako
T
Snellman
Anne
A
Göhring
Walter
W
Pirilä
Paivi
P
Timpl
Rupert
R
Pihlajaniemi
Taina
T
D000595:Amino Acid Sequence; D001665:Binding Sites; D002135:Calcium-Binding Proteins; D002352:Carrier Proteins / Q000737:chemistry*; D015815:Cell Adhesion Molecules; D002942:Circular Dichroism; D003094:Collagen / Q000737:chemistry; D024181:Collagen Type XIII / Q000737:chemistry*; D019281:Dimerization; D004305:Dose-Response Relationship, Drug; D005353:Fibronectins / Q000737:chemistry*; D006023:Glycoproteins / Q000378:metabolism; D019812:Heparan Sulfate Proteoglycans / Q000737:chemistry*; D006493:Heparin / Q000737:chemistry*; D006801:Humans; D007700:Kinetics; D008562:Membrane Glycoproteins / Q000737:chemistry*; D008854:Microscopy, Electron; D008954:Models, Biological; D008969:Molecular Sequence Data; D010434:Pepsin A / Q000737:chemistry; D011485:Protein Binding; D017434:Protein Structure, Tertiary; D011994:Recombinant Proteins / Q000378:metabolism; D020349:Surface Plasmon Resonance; D013696:Temperature; D013997:Time Factors
D002135:Calcium-Binding Proteins; D002352:Carrier Proteins; D015815:Cell Adhesion Molecules; D024181:Collagen Type XIII; D005353:Fibronectins; D006023:Glycoproteins; D019812:Heparan Sulfate Proteoglycans; D008562:Membrane Glycoproteins; C114729:NID2 protein, human; D011994:Recombinant Proteins; C071927:perlecan; D006493:Heparin; D003094:Collagen; D010434:Pepsin A