Variable zinc coordination in endostatin.
Endostatin is a proteolytic fragment of collagen XVIII that potently inhibits angiogenesis and tumour growth. Human endostatin contains a zinc ion, bound near the N terminus, which was not observed in the original structure of mouse endostatin at pH 5. Controversial data exist on the role of this zinc ion in the anti-tumour activity. We report two new crystal structures of mouse endostatin at pH 8.5 with bound zinc. One crystal form shows a metal ion coordination similar to that in human endostatin (His132, His134, His142, Asp207), but the conformation of the N-terminal segment is different. In the other crystal form, Asp136 replaces His132 as a zinc ligand. Site-directed mutagenesis of zinc-binding residues demonstrates that both coordination geometries occur in solution. The large degree of structural heterogeneity of the zinc-binding site has implications for endostatin function. We conclude that zinc is likely to play a structural rather than a critical functional role in endostatin.
Hohenester
E
E
Biophysics Section, Blackett Laboratory, Imperial College, London, SW7 2AZ, UK. hohenester@ic.ac.uk
Sasaki
T
T
Mann
K
K
Timpl
R
R
D000595:Amino Acid Sequence; D019943:Amino Acid Substitution; D000818:Animals; D001224:Aspartic Acid / Q000235:genetics / Q000378:metabolism; D001665:Binding Sites; D003094:Collagen / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism*; D043170:Collagen Type XVIII; D018360:Crystallography, X-Ray; D019281:Dimerization; D043169:Endostatins; D006639:Histidine / Q000235:genetics / Q000378:metabolism; D006801:Humans; D051379:Mice; D008958:Models, Molecular; D008969:Molecular Sequence Data; D010446:Peptide Fragments / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism*; D011487:Protein Conformation; D012996:Solutions; D013329:Structure-Activity Relationship; D015032:Zinc / Q000378:metabolism*
D043170:Collagen Type XVIII; D043169:Endostatins; D010446:Peptide Fragments; D012996:Solutions; D001224:Aspartic Acid; D006639:Histidine; D003094:Collagen; D015032:Zinc