title: Inhibition of the cysteine proteinases cathepsins K and L by the serpin headpin (SERPINB13): a kinetic analysis.
abstract: Headpin (SERPINB13) is a novel member of the serine proteinase inhibitor (Serpin) gene family that was originally cloned from a keratinocyte cDNA library. Western blot analysis using a headpin-specific antiserum recognized a protein with the predicted M(r) of 44kDa in lysates derived from a transformed keratinocyte cell line known to express headpin mRNA. Similarity of the reactive-site loop (RSL) domain of headpin, notably at the P1-P1(') residues, with other serpins that inhibit cysteine and serine proteinases suggests that headpin may inhibit similar proteinases. This study demonstrates that recombinant headpin indeed inhibits cathepsins K and L, but not chymotrypsin, elastase, trypsin, subtilisin A, urokinase-type plasminogen activator, plasmin, or thrombin. The second-order rate constants (k(a)) for the inhibitory reactions of rHeadpin with cathepsins K and L were 5.1+/-0.6x10(4) and 4.1+/-0.8x10(4)M(-1)s(-1), respectively. Headpin formed SDS-stable complexes with cathepsins K and L, a characteristic property of inhibitory serpins. Interactions of the RSL domain of headpin with cathepsins K and L were indicated by cleavage of headpin near the predicted P1-P1(') residues by these proteinases. These results demonstrate that the serpin headpin possesses specificity for inhibiting lysosomal cysteine proteinases.
authors.affiliation: Department of Head and Neck Surgery, M.D. Anderson Cancer Center, The University of Texas, 1515 Holcombe Blvd., Houston, TX 77030-4095, USA.
authors.forename: Arumugam
authors.initials: A
authors.lastname: Jayakumar
authors.forename: Ya'an
authors.initials: Y
authors.lastname: Kang
authors.forename: Mitchell J
authors.initials: MJ
authors.lastname: Frederick
authors.forename: Stephen C
authors.initials: SC
authors.lastname: Pak
authors.forename: Ying
authors.initials: Y
authors.lastname: Henderson
authors.forename: Paula R
authors.initials: PR
authors.lastname: Holton
authors.forename: Kenji
authors.initials: K
authors.lastname: Mitsudo
authors.forename: Gary A
authors.initials: GA
authors.lastname: Silverman
authors.forename: Adel K
authors.initials: AK
authors.lastname: EL-Naggar
authors.forename: Dieter
authors.initials: D
authors.lastname: Brömme
authors.forename: Gary L
authors.initials: GL
authors.lastname: Clayman
mesh_terms: D000595:Amino Acid Sequence; D000818:Animals; D016367:Baculoviridae / Q000235:genetics; D056657:Cathepsin K; D056668:Cathepsin L; D002403:Cathepsins / Q000037:antagonists & inhibitors* / Q000187:drug effects / Q000378:metabolism; D002461:Cell Line, Transformed; D003546:Cysteine Endopeptidases / Q000187:drug effects / Q000378:metabolism*; D006801:Humans; D015603:Keratinocytes / Q000166:cytology; D007700:Kinetics; D008247:Lysosomes / Q000201:enzymology; D008969:Molecular Sequence Data; D008970:Molecular Weight; D017434:Protein Structure, Tertiary; D012333:RNA, Messenger / Q000378:metabolism; D011993:Recombinant Fusion Proteins / Q000494:pharmacology; D015842:Serine Proteinase Inhibitors / Q000494:pharmacology*; D015843:Serpins / Q000737:chemistry / Q000302:isolation & purification / Q000494:pharmacology*; D012967:Sodium Dodecyl Sulfate / Q000378:metabolism; D019032:Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization; D018411:Spodoptera / Q000166:cytology / Q000235:genetics / Q000821:virology
chemical_list: D012333:RNA, Messenger; D011993:Recombinant Fusion Proteins; C530652:SERPINB13 protein, human; D015842:Serine Proteinase Inhibitors; D015843:Serpins; D012967:Sodium Dodecyl Sulfate; D002403:Cathepsins; D003546:Cysteine Endopeptidases; C534281:CTSL protein, human; D056668:Cathepsin L; C534130:CTSK protein, human; D056657:Cathepsin K