The action of neutrophil serine proteases on elastin and its precursor.
This study aimed to investigate the degradation of the natural substrates tropoelastin and elastin by the neutrophil-derived serine proteases human leukocyte elastase (HLE), proteinase 3 (PR3) and cathepsin G (CG). Focus was placed on determining their cleavage site specificities using mass spectrometric techniques. Moreover, the release of bioactive peptides from elastin by the three proteases was studied. Tropoelastin was comprehensively degraded by all three proteases, whereas less cleavage occurred in mature cross-linked elastin. An analysis of the cleavage site specificities of the three proteases in tropoelastin and elastin revealed that HLE and PR3 similarly tolerate hydrophobic and/or aliphatic amino acids such as Ala, Gly and Val at P(1), which are also preferred by CG. In addition, CG prefers the bulky hydrophobic amino acid Leu and accepts the bulky aromatic amino acids Phe and Tyr. CG shows a strong preference for the charged amino acid Lys at P(1) in tropoelastin, whereas Lys was not identified at P(1) in CG digests of elastin due to extensive cross-linking at Lys residues in mature elastin. All three serine proteases showed a clear preference for Pro at P(2) and P(4)'. With respect to the liberation of potentially bioactive peptides from elastin, the study revealed that all three serine proteases have a similar ability to release bioactive sequences, with CG producing the highest number of these peptides. In bioactivity studies, potentially bioactive peptides that have not been investigated on their bioactivity to date, were tested. Three new bioactive GxxPG motifs were identified; GVYPG, GFGPG and GVLPG.
Institute of Pharmacy, Martin Luther University Halle-Wittenberg, Faculty of Natural Sciences I, Wolfgang-Langenbeck-Str. 4, 06120 Halle (Saale), Germany.
Andrea
A
Heinz
Michael C
MC
Jung
Günther
G
Jahreis
Anthony
A
Rusciani
Laurent
L
Duca
Laurent
L
Debelle
Anthony S
AS
Weiss
Reinhard H H
RH
Neubert
Christian E H
CE
Schmelzer
D000595:Amino Acid Sequence; D002851:Chromatography, High Pressure Liquid; D004549:Elastin / Q000378:metabolism*; D004797:Enzyme-Linked Immunosorbent Assay; D006801:Humans; D019272:Leukocyte Elastase / Q000378:metabolism*; D008969:Molecular Sequence Data; D009504:Neutrophils / Q000201:enzymology*; D011498:Protein Precursors / Q000378:metabolism*; D019032:Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization
D011498:Protein Precursors; D004549:Elastin; D019272:Leukocyte Elastase