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Human Glandular Kallikrein 2

Lövgren, Janita LU (1999)
Abstract
Human glandular kallikrein 2 (hK2) is a serine protease, which is predominantly expressed by the prostate gland and found in seminal plasma at mean levels of 6 mg/ml. It has 79% amino acid sequence identity with prostate-specific antigen (PSA or hK3).



The aim of this work was to produce hK2 by recombinant expression technologies in order to study its biochemical and biological properties and its cross-reaction with PSA immunoassays. Recombinant hK2 was produced in parallel with PSA in mammalian and insect cells. The two proteins were purified, characterized and shown to be immunologically cross-reactive. Recombinant PSA was expressed at high levels and recovered as an inactive zymogen (proPSA). The expression levels of... (More)
Human glandular kallikrein 2 (hK2) is a serine protease, which is predominantly expressed by the prostate gland and found in seminal plasma at mean levels of 6 mg/ml. It has 79% amino acid sequence identity with prostate-specific antigen (PSA or hK3).



The aim of this work was to produce hK2 by recombinant expression technologies in order to study its biochemical and biological properties and its cross-reaction with PSA immunoassays. Recombinant hK2 was produced in parallel with PSA in mammalian and insect cells. The two proteins were purified, characterized and shown to be immunologically cross-reactive. Recombinant PSA was expressed at high levels and recovered as an inactive zymogen (proPSA). The expression levels of recombinant hK2 were low and hK2 was always recovered in an enzymatically active mature form. Mutation of the propeptide of hK2 resulted in recovery of hK2 as an inactive zymogen, and a 15 to 40 fold increase in the expression levels.



We have shown that proPSA can be activated by hK2 at physiological PSA to hK2 ratios, making hK2 a likely activator of PSA in prostatic fluid. PSA purified from the spent medium of LNCaP cells, which produce both PSA and hK2, was only partially converted into mature form. This could be due to a slow activation process in culture medium. HK2 cleaves the gel-proteins semenogelin I and II, and tripeptide substrates on the carboxy-terminal side of certain single and double arginines.



We also studied the regulation of hK2 by several extracellular protease inhibitors of which only protein C inhibitor, PCI, was able to rapidly inhibit hK2. Zinc is present at extremely high concentrations in the prostate, and zinc ions were shown to be able to inhibit hK2 at micromolar levels. (Less)
Please use this url to cite or link to this publication:
author
opponent
  • PhD Diamandis, Eleftherios P, Head, Sect. Clin. Biochem., Dept. Path. and Lab. Med., Mount Sinai Hospital, and Professor and Head, Div. Clinical Biochemistry, Dept. Laboratory Medicine and Pathobiology, University of Toronto, Canada
organization
publishing date
type
Thesis
publication status
published
subject
keywords
seminal plasma, semenogelin, PCI, serpin, zinc, cancer, prostate, PSA, hK2, serine protease, kallikrein, Clinical chemistry, Klinisk kemi
pages
128 pages
publisher
Janita Lövgren, Munkkentie 8 E 5, 21620 Kuusisto, Finland,
defense location
Lilla Aulan, Medicinskt Forskningscentrum, Malmö
defense date
1999-12-16 10:15
external identifiers
  • Other:ISRN: LUMEDW/MECM-1023-SE
ISBN
91-628-3891-1
language
English
LU publication?
yes
id
8dd607d3-8197-4791-be6e-bd67937c2159 (old id 40058)
date added to LUP
2007-07-31 13:25:41
date last changed
2016-09-19 08:45:06
@misc{8dd607d3-8197-4791-be6e-bd67937c2159,
  abstract     = {Human glandular kallikrein 2 (hK2) is a serine protease, which is predominantly expressed by the prostate gland and found in seminal plasma at mean levels of 6 mg/ml. It has 79% amino acid sequence identity with prostate-specific antigen (PSA or hK3).<br/><br>
<br/><br>
The aim of this work was to produce hK2 by recombinant expression technologies in order to study its biochemical and biological properties and its cross-reaction with PSA immunoassays. Recombinant hK2 was produced in parallel with PSA in mammalian and insect cells. The two proteins were purified, characterized and shown to be immunologically cross-reactive. Recombinant PSA was expressed at high levels and recovered as an inactive zymogen (proPSA). The expression levels of recombinant hK2 were low and hK2 was always recovered in an enzymatically active mature form. Mutation of the propeptide of hK2 resulted in recovery of hK2 as an inactive zymogen, and a 15 to 40 fold increase in the expression levels.<br/><br>
<br/><br>
We have shown that proPSA can be activated by hK2 at physiological PSA to hK2 ratios, making hK2 a likely activator of PSA in prostatic fluid. PSA purified from the spent medium of LNCaP cells, which produce both PSA and hK2, was only partially converted into mature form. This could be due to a slow activation process in culture medium. HK2 cleaves the gel-proteins semenogelin I and II, and tripeptide substrates on the carboxy-terminal side of certain single and double arginines.<br/><br>
<br/><br>
We also studied the regulation of hK2 by several extracellular protease inhibitors of which only protein C inhibitor, PCI, was able to rapidly inhibit hK2. Zinc is present at extremely high concentrations in the prostate, and zinc ions were shown to be able to inhibit hK2 at micromolar levels.},
  author       = {Lövgren, Janita},
  isbn         = {91-628-3891-1},
  keyword      = {seminal plasma,semenogelin,PCI,serpin,zinc,cancer,prostate,PSA,hK2,serine protease,kallikrein,Clinical chemistry,Klinisk kemi},
  language     = {eng},
  pages        = {128},
  publisher    = {ARRAY(0x8ced9c0)},
  title        = {Human Glandular Kallikrein 2},
  year         = {1999},
}