Inhibition of serine proteinases plasmin, trypsin, subtilisin A, cathepsin G, and elastase by LEKTI: a kinetic analysis.
The human LEKTI gene encodes a putative 15-domain serine proteinase inhibitor and has been linked to the inherited disorder known as Netherton syndrome. In this study, human recombinant LEKTI (rLEKTI) was purified using a baculovirus/insect cell expression system, and the inhibitory profile of the full-length rLEKTI protein was examined. Expression of LEKTI in Sf9 cells showed the presence of disulfide bonds, suggesting the maintenance of the tertiary protein structure. rLEKTI inhibited the serine proteinases plasmin, subtilisin A, cathepsin G, human neutrophil elastase, and trypsin, but not chymotrypsin. Moreover, rLEKTI did not inhibit the cysteine proteinase papain or cathepsin K, L, or S. Further, rLEKTI inhibitory activity was inactivated by treatment with 20 mM DTT, suggesting that disulfide bonds are important to LEKTI function. The inhibition of plasmin, subtilisin A, cathepsin G, elastase, and trypsin by rLEKTI occurred through a noncompetitive-type mechanism, with inhibitory constants (K(i)) of 27 +/- 5, 49 +/- 3, 67 +/- 6, 317 +/-36, and 849 +/- 55 nM, respectively. Thus, LEKTI is likely to be a major physiological inhibitor of multiple serine proteinases.
Mitsudo
Kenji
K
Department of Head and Neck Surgery, The University of Texas M. D. Anderson Cancer Center, 1515 Holcombe Boulevard, Houston, Texas 77030-4095, USA.
Jayakumar
Arumugam
A
Henderson
Ying
Y
Frederick
Mitchell J
MJ
Kang
Ya'an
Y
Wang
Mary
M
El-Naggar
Adel K
AK
Clayman
Gary L
GL
D000595:Amino Acid Sequence; D000818:Animals; D016367:Baculoviridae / Q000235:genetics; D001483:Base Sequence; D002352:Carrier Proteins; D056649:Cathepsin G; D002403:Cathepsins / Q000037:antagonists & inhibitors*; D002461:Cell Line, Transformed; D002918:Chymotrypsin / Q000378:metabolism; D017931:DNA Primers / Q000737:chemistry; D018076:DNA, Complementary / Q000235:genetics / Q000378:metabolism; D004220:Disulfides / Q000737:chemistry*; D004229:Dithiothreitol / Q000378:metabolism; D005341:Fibrinolysin / Q000037:antagonists & inhibitors*; D006801:Humans; D008969:Molecular Sequence Data; D010084:Oxidation-Reduction; D010196:Pancreatic Elastase / Q000037:antagonists & inhibitors*; D010940:Plant Proteins / Q000494:pharmacology*; D016133:Polymerase Chain Reaction; D053491:Proteinase Inhibitory Proteins, Secretory; D011993:Recombinant Fusion Proteins / Q000494:pharmacology; D012697:Serine Endopeptidases; D000074163:Serine Peptidase Inhibitor Kazal-Type 5; D015842:Serine Proteinase Inhibitors / Q000235:genetics / Q000494:pharmacology*; D018411:Spodoptera / Q000166:cytology / Q000235:genetics / Q000821:virology; D013381:Subtilisins / Q000037:antagonists & inhibitors*; D014361:Trypsin Inhibitors; D000516:alpha-Amylases / Q000037:antagonists & inhibitors
D002352:Carrier Proteins; D017931:DNA Primers; D018076:DNA, Complementary; D004220:Disulfides; D010940:Plant Proteins; D053491:Proteinase Inhibitory Proteins, Secretory; D011993:Recombinant Fusion Proteins; C120650:SPINK5 protein, human; D000074163:Serine Peptidase Inhibitor Kazal-Type 5; D015842:Serine Proteinase Inhibitors; D014361:Trypsin Inhibitors; D000516:alpha-Amylases; D002403:Cathepsins; D012697:Serine Endopeptidases; D013381:Subtilisins; D002918:Chymotrypsin; C534094:CTSG protein, human; D056649:Cathepsin G; D010196:Pancreatic Elastase; D005341:Fibrinolysin; D004229:Dithiothreitol