Purification and characterization of human kallikrein 11, a candidate prostate and ovarian cancer biomarker, from seminal plasma

Liu Ying Luo, Shannon J C Shan, Marc B. Elliott, Antoninus Soosaipillai, Eleftherios P. Diamandis

Research output: Contribution to journalArticle

Abstract

Purpose: Preliminary data suggest that hK11 is a novel serum biomarker for prostate and ovarian cancer. To examine the enzymatic characteristics of hK11, we purified and functionally characterized native hK11 from seminal plasma. Experimental Design: hK11 was purified from seminal plasma by immunoaffinity chromatography and characterized by kinetic analysis, electrophoresis, Western blots, and mass spectrometry. Results: hK11 is present in seminal plasma at concentrations ranging from 2 to 37 μg/mL. Using immunoaffinity chromatography and reverse-phase high-performance liquid chromatography, we purified hK11 to homogeneity. In seminal plasma, hK11 is present as a free enzyme of ∼40 kDa. About 40% of hK11 is enzymaticalty active, whereas the rest is inactivated by internal cleavage after Arg156 (Genbank accession no. AF164623), which generates two peptides of ∼20 kDa, connected by internal disulfide bonds. Purified hK11 possesses trypsin-like activity and cleaves synthetic peptides after arginine but not lysine residues. It does not cleave chymotrypsin substrates. Antithrombin, α1- antichymotrypsin, α2-antiplasmin, and α1- antitrypsin have no effect on hK11 activity and do not form complexes with hK11 in vitro. The strongest inhibitor, APMSF, completely inhibited hK11 activity at a concentration of 2.5 mmol/L. Aprotinin and an hK11-specific monoclonal antibody inhibited hK11 activity up to 40%. Plasmin is a strong candidate for cleaving hK11 at Arg156. Conclusion: This is the first report on purification and characterization of native hK11. We speculate that hK11, along with other kallikreins, proteases, and inhibitors, participates in a cascade enzymatic pathway responsible for semen liquefaction after ejaculation.

Original languageEnglish (US)
Pages (from-to)742-750
Number of pages9
JournalClinical Cancer Research
Volume12
Issue number3 I
DOIs
StatePublished - Feb 1 2006
Externally publishedYes

Fingerprint

Kallikreins
Tumor Biomarkers
Semen
Ovarian Neoplasms
Prostatic Neoplasms
Chromatography
Peptides
Antifibrinolytic Agents
Ejaculation
Aprotinin
Antithrombins
Fibrinolysin
Nucleic Acid Databases
Chymotrypsin
Reverse-Phase Chromatography
Protease Inhibitors
Disulfides
Trypsin
Lysine
Arginine

ASJC Scopus subject areas

  • Cancer Research
  • Oncology

Cite this

Purification and characterization of human kallikrein 11, a candidate prostate and ovarian cancer biomarker, from seminal plasma. / Luo, Liu Ying; Shan, Shannon J C; Elliott, Marc B.; Soosaipillai, Antoninus; Diamandis, Eleftherios P.

In: Clinical Cancer Research, Vol. 12, No. 3 I, 01.02.2006, p. 742-750.

Research output: Contribution to journalArticle

Luo, Liu Ying ; Shan, Shannon J C ; Elliott, Marc B. ; Soosaipillai, Antoninus ; Diamandis, Eleftherios P. / Purification and characterization of human kallikrein 11, a candidate prostate and ovarian cancer biomarker, from seminal plasma. In: Clinical Cancer Research. 2006 ; Vol. 12, No. 3 I. pp. 742-750.
@article{94d0e929d7584f7697f29fce014142e7,
title = "Purification and characterization of human kallikrein 11, a candidate prostate and ovarian cancer biomarker, from seminal plasma",
abstract = "Purpose: Preliminary data suggest that hK11 is a novel serum biomarker for prostate and ovarian cancer. To examine the enzymatic characteristics of hK11, we purified and functionally characterized native hK11 from seminal plasma. Experimental Design: hK11 was purified from seminal plasma by immunoaffinity chromatography and characterized by kinetic analysis, electrophoresis, Western blots, and mass spectrometry. Results: hK11 is present in seminal plasma at concentrations ranging from 2 to 37 μg/mL. Using immunoaffinity chromatography and reverse-phase high-performance liquid chromatography, we purified hK11 to homogeneity. In seminal plasma, hK11 is present as a free enzyme of ∼40 kDa. About 40{\%} of hK11 is enzymaticalty active, whereas the rest is inactivated by internal cleavage after Arg156 (Genbank accession no. AF164623), which generates two peptides of ∼20 kDa, connected by internal disulfide bonds. Purified hK11 possesses trypsin-like activity and cleaves synthetic peptides after arginine but not lysine residues. It does not cleave chymotrypsin substrates. Antithrombin, α1- antichymotrypsin, α2-antiplasmin, and α1- antitrypsin have no effect on hK11 activity and do not form complexes with hK11 in vitro. The strongest inhibitor, APMSF, completely inhibited hK11 activity at a concentration of 2.5 mmol/L. Aprotinin and an hK11-specific monoclonal antibody inhibited hK11 activity up to 40{\%}. Plasmin is a strong candidate for cleaving hK11 at Arg156. Conclusion: This is the first report on purification and characterization of native hK11. We speculate that hK11, along with other kallikreins, proteases, and inhibitors, participates in a cascade enzymatic pathway responsible for semen liquefaction after ejaculation.",
author = "Luo, {Liu Ying} and Shan, {Shannon J C} and Elliott, {Marc B.} and Antoninus Soosaipillai and Diamandis, {Eleftherios P.}",
year = "2006",
month = "2",
day = "1",
doi = "10.1158/1078-0432.CCR-05-1696",
language = "English (US)",
volume = "12",
pages = "742--750",
journal = "Clinical Cancer Research",
issn = "1078-0432",
publisher = "American Association for Cancer Research Inc.",
number = "3 I",

}

TY - JOUR

T1 - Purification and characterization of human kallikrein 11, a candidate prostate and ovarian cancer biomarker, from seminal plasma

AU - Luo, Liu Ying

AU - Shan, Shannon J C

AU - Elliott, Marc B.

AU - Soosaipillai, Antoninus

AU - Diamandis, Eleftherios P.

PY - 2006/2/1

Y1 - 2006/2/1

N2 - Purpose: Preliminary data suggest that hK11 is a novel serum biomarker for prostate and ovarian cancer. To examine the enzymatic characteristics of hK11, we purified and functionally characterized native hK11 from seminal plasma. Experimental Design: hK11 was purified from seminal plasma by immunoaffinity chromatography and characterized by kinetic analysis, electrophoresis, Western blots, and mass spectrometry. Results: hK11 is present in seminal plasma at concentrations ranging from 2 to 37 μg/mL. Using immunoaffinity chromatography and reverse-phase high-performance liquid chromatography, we purified hK11 to homogeneity. In seminal plasma, hK11 is present as a free enzyme of ∼40 kDa. About 40% of hK11 is enzymaticalty active, whereas the rest is inactivated by internal cleavage after Arg156 (Genbank accession no. AF164623), which generates two peptides of ∼20 kDa, connected by internal disulfide bonds. Purified hK11 possesses trypsin-like activity and cleaves synthetic peptides after arginine but not lysine residues. It does not cleave chymotrypsin substrates. Antithrombin, α1- antichymotrypsin, α2-antiplasmin, and α1- antitrypsin have no effect on hK11 activity and do not form complexes with hK11 in vitro. The strongest inhibitor, APMSF, completely inhibited hK11 activity at a concentration of 2.5 mmol/L. Aprotinin and an hK11-specific monoclonal antibody inhibited hK11 activity up to 40%. Plasmin is a strong candidate for cleaving hK11 at Arg156. Conclusion: This is the first report on purification and characterization of native hK11. We speculate that hK11, along with other kallikreins, proteases, and inhibitors, participates in a cascade enzymatic pathway responsible for semen liquefaction after ejaculation.

AB - Purpose: Preliminary data suggest that hK11 is a novel serum biomarker for prostate and ovarian cancer. To examine the enzymatic characteristics of hK11, we purified and functionally characterized native hK11 from seminal plasma. Experimental Design: hK11 was purified from seminal plasma by immunoaffinity chromatography and characterized by kinetic analysis, electrophoresis, Western blots, and mass spectrometry. Results: hK11 is present in seminal plasma at concentrations ranging from 2 to 37 μg/mL. Using immunoaffinity chromatography and reverse-phase high-performance liquid chromatography, we purified hK11 to homogeneity. In seminal plasma, hK11 is present as a free enzyme of ∼40 kDa. About 40% of hK11 is enzymaticalty active, whereas the rest is inactivated by internal cleavage after Arg156 (Genbank accession no. AF164623), which generates two peptides of ∼20 kDa, connected by internal disulfide bonds. Purified hK11 possesses trypsin-like activity and cleaves synthetic peptides after arginine but not lysine residues. It does not cleave chymotrypsin substrates. Antithrombin, α1- antichymotrypsin, α2-antiplasmin, and α1- antitrypsin have no effect on hK11 activity and do not form complexes with hK11 in vitro. The strongest inhibitor, APMSF, completely inhibited hK11 activity at a concentration of 2.5 mmol/L. Aprotinin and an hK11-specific monoclonal antibody inhibited hK11 activity up to 40%. Plasmin is a strong candidate for cleaving hK11 at Arg156. Conclusion: This is the first report on purification and characterization of native hK11. We speculate that hK11, along with other kallikreins, proteases, and inhibitors, participates in a cascade enzymatic pathway responsible for semen liquefaction after ejaculation.

UR - http://www.scopus.com/inward/record.url?scp=32944456614&partnerID=8YFLogxK

UR - http://www.scopus.com/inward/citedby.url?scp=32944456614&partnerID=8YFLogxK

U2 - 10.1158/1078-0432.CCR-05-1696

DO - 10.1158/1078-0432.CCR-05-1696

M3 - Article

C2 - 16467084

AN - SCOPUS:32944456614

VL - 12

SP - 742

EP - 750

JO - Clinical Cancer Research

JF - Clinical Cancer Research

SN - 1078-0432

IS - 3 I

ER -