Structural characterisation of human stefin A in solution and implications for binding to cysteine proteinases.
Stefin A is a member of the cystatin superfamily of proteins which are tight and reversibly binding inhibitors of the papain-like cysteine proteinases. The 1H-NMR and 15N-NMR resonances of human stefin A have been sequentially assigned using two-dimensional homonuclear and heteronuclear NMR techniques in conjunction with three-dimensional heteronuclear methods. Characteristic sequential and medium range NOE contacts, J constants and hydrogen exchange data have been used to identify the secondary structural elements of the protein which consists of five anti-parallel beta-strands and a single alpha-helix. There is much similarity between the secondary structural features of stefin A and the homologous protein stefin B in its complex with papain [Stubbs, M. T., Laber, B., Bode, W., Huber, R., Jerala, R., Lenarcic, B. & Turk, V. (1990) EMBO. J. 9. 1939-1947] but also some important differences in regions which are fundamental to the binding event. The principal difference is the presence of two conformationally unrestricted regions in stefin A that form two of the components of the tripartite wedge which docks into the active site of the target proteinase. Specifically, these regions are the five N-terminal residues and the second binding loop, which form a turn and a short helix respectively, in the bound conformation of stefin B.
Martin
J R
JR
Krebs Institute, Department of Molecular Biology and Biotechnology, University of Sheffield, England.
Jerala
R
R
Kroon-Zitko
L
L
Zerovnik
E
E
Turk
V
V
Waltho
J P
JP
D000595:Amino Acid Sequence; D001665:Binding Sites; D055312:Cystatin A; D055313:Cystatin B; D015891:Cystatins / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism*; D003546:Cysteine Endopeptidases / Q000378:metabolism*; D004578:Electron Spin Resonance Spectroscopy; D006801:Humans; D009682:Magnetic Resonance Spectroscopy; D008969:Molecular Sequence Data; D015394:Molecular Structure; D010449:Peptide Mapping; D011487:Protein Conformation; D017433:Protein Structure, Secondary; D011994:Recombinant Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D012996:Solutions
C524994:CSTB protein, human; D055312:Cystatin A; D015891:Cystatins; D011994:Recombinant Proteins; D012996:Solutions; C026407:CSTA protein, human; D055313:Cystatin B; D003546:Cysteine Endopeptidases