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Table 3 Oligopeptidase activities in the culture supernatant, cell lysate and membrane fraction of B16F10-Nex2 cells

From: Characterization of thimet oligopeptidase and neurolysin activities in B16F10-Nex2 tumor cells and their involvement in angiogenesis and tumor growth

Cell fraction

Specific activity (μM/min per mg)

Activity per cell (pM/min per 103 cells)

Total activity (μM/min)

Supernatant

1.39 ± 0.24

36 ± 16

3.36 ± 0.15

Lysate

3.22 ± 0.96

326 ± 15

31.56 ± 0.16

Lysate + Anti-TOP Ab

0.64 ± 0.19

65 ± 5

6.31 ± 0.03

Lysate + Anti-Neurolysin Ab

2.58 ± 0.77

261 ± 5

25.25 ± 0.13

Membrane*

< 0.01

3 ± 0.5

0.2 ± 0.01

  1. Culture supernatant, cell lysate and membrane preparation of B16F10-Nex2 cells were incubated with the fluorogenic substrate Abz-GFSPFRQ-EDDnp (20 μM) in 50 mM Tris-HCl, pH 7.4 at 37°C. Enzyme activity was followed by measuring the fluorescence at λem. = 420 nm and λex. = 320 nm. Results are expressed as means ± SD. Ab= antibodies, used at 1:300.
  2. *Although the activity of TOP in the membrane was low, the protein was clearly detected by Western blotting with anti-TOP antibody.
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Molecular Cancer

ISSN: 1476-4598