Degradation of tropoelastin by matrix metalloproteinases--cleavage site specificities and release of matrikines

TY - JOUR

T1 - Degradation of tropoelastin by matrix metalloproteinases--cleavage site specificities and release of matrikines

AU - Heinz, Andrea

AU - Jung, Michael C

AU - Duca, Laurent

AU - Sippl, Wolfgang

AU - Taddese, Samuel

AU - Ihling, Christian

AU - Rusciani, Anthony

AU - Jahreis, Günther

AU - Weiss, Anthony S

AU - Neubert, Reinhard H H

AU - Schmelzer, Christian E H

PY - 2010/4

Y1 - 2010/4

N2 - To provide a basis for the development of approaches to treat elastin-degrading diseases, the aim of this study was to investigate the degradation of the natural substrate tropoelastin by the elastinolytic matrix metalloproteinases MMP-7, MMP-9, and MMP-12 and to compare the cleavage site specificities of the enzymes using complementary MS techniques and molecular modeling. Furthermore, the ability of the three proteases to release bioactive peptides was studied. Tropoelastin was readily degraded by all three MMPs. Eighty-nine cleavage sites in tropoelastin were identified for MMP-12, whereas MMP-7 and MMP-9 were found to cleave at only 58 and 63 sites, respectively. Cleavages occurred predominantly in the N-terminal and C-terminal regions of tropoelastin. With respect to the cleavage site specificities, the study revealed that all three MMPs similarly tolerate hydrophobic and/or aliphatic amino acids, including Pro, Gly, Ile, and Val, at P(1)'. MMP-7 shows a strong preference for Leu at P(1)', which is also well accepted by MMP-9 and MMP-12. Of all three MMPs, MMP-12 best tolerates bulky charged and aromatic amino acids at P(1)'. All three MMPs showed a clear preference for Pro at P(3) that could be structurally explained by molecular modeling. Analysis of the generated peptides revealed that all three MMPs show a similar ability to release bioactive sequences, with MMP-12 producing the highest number of these peptides. Furthermore, the generated peptides YTTGKLPYGYGPGG, YGARPGVGVGGIP, and PGFGAVPGA, containing GxxPG motifs that have not yet been proven to be bioactive, were identified as new matrikines upon biological activity testing.

AB - To provide a basis for the development of approaches to treat elastin-degrading diseases, the aim of this study was to investigate the degradation of the natural substrate tropoelastin by the elastinolytic matrix metalloproteinases MMP-7, MMP-9, and MMP-12 and to compare the cleavage site specificities of the enzymes using complementary MS techniques and molecular modeling. Furthermore, the ability of the three proteases to release bioactive peptides was studied. Tropoelastin was readily degraded by all three MMPs. Eighty-nine cleavage sites in tropoelastin were identified for MMP-12, whereas MMP-7 and MMP-9 were found to cleave at only 58 and 63 sites, respectively. Cleavages occurred predominantly in the N-terminal and C-terminal regions of tropoelastin. With respect to the cleavage site specificities, the study revealed that all three MMPs similarly tolerate hydrophobic and/or aliphatic amino acids, including Pro, Gly, Ile, and Val, at P(1)'. MMP-7 shows a strong preference for Leu at P(1)', which is also well accepted by MMP-9 and MMP-12. Of all three MMPs, MMP-12 best tolerates bulky charged and aromatic amino acids at P(1)'. All three MMPs showed a clear preference for Pro at P(3) that could be structurally explained by molecular modeling. Analysis of the generated peptides revealed that all three MMPs show a similar ability to release bioactive sequences, with MMP-12 producing the highest number of these peptides. Furthermore, the generated peptides YTTGKLPYGYGPGG, YGARPGVGVGGIP, and PGFGAVPGA, containing GxxPG motifs that have not yet been proven to be bioactive, were identified as new matrikines upon biological activity testing.

KW - Amino Acid Motifs

KW - Amino Acid Sequence

KW - Binding Sites

KW - Humans

KW - Mass Spectrometry

KW - Matrix Metalloproteinase 12

KW - Matrix Metalloproteinase 7

KW - Matrix Metalloproteinase 9

KW - Matrix Metalloproteinases

KW - Models, Molecular

KW - Molecular Sequence Data

KW - Molecular Weight

KW - Peptides

KW - Protein Binding

KW - Protein Structure, Tertiary

KW - Recombinant Proteins

KW - Substrate Specificity

KW - Tropoelastin

KW - Journal Article

KW - Research Support, Non-U.S. Gov't

U2 - 10.1111/j.1742-4658.2010.07616.x

DO - 10.1111/j.1742-4658.2010.07616.x

M3 - Journal article

C2 - 20345904

SN - 1742-464X

VL - 277

SP - 1939

EP - 1956

JO - F E B S Journal

JF - F E B S Journal

IS - 8

ER -