MASP-1 and MASP-2 do not activate pro-factor D in resting human blood, whereas MASP-3 is a potential activator: Kinetic analysis involving specific MASP-1 and MASP-2 inhibitors

Gábor Oroszlán, Elod Kortvely, Dávid Szakács, Andrea Kocsis, Sascha Dammeier, Anne Zeck, Marius Ueffing, P. Závodszky, G. Pál, P. Gál, J. Dobó

Research output: Contribution to journalArticle

22 Citations (Scopus)

Abstract

It had been thought that complement factor D (FD) is activated at the site of synthesis, and only FD lacking a propeptide is present in blood. The serum of mannose-binding lectin-associated serine protease (MASP)-1/3(2/2) mice contains pro-FD and has markedly reduced alternative pathway activity. It was suggested that MASP-1 and MASP-3 directly activate pro-FD; however, other experiments contradicted this view. We decided to clarify the involvement of MASPs in pro-FD activation in normal, as opposed to deficient, human plasma and serum. Human pro-FD containing an APPRGR propeptide was produced in insect cells. We measured its activation kinetics using purified active MASP-1, MASP-2, MASP-3, as well as thrombin.We found all these enzymes to be efficient activators, whereas MASP proenzymes lacked such activity. Pro-FD cleavage in serum or plasma was quantified by a novel assay using fluorescently labeled pro-FD. Labeled pro-FD was processed with t1/2s of ∼3 and 5 h in serum and plasma, respectively, showing that proteolytic activity capable of activating pro-FD exists in blood even in the absence of active coagulation enzymes. Our previously developed selective MASP-1 and MASP-2 inhibitors did not reduce pro-FD activation at reasonable concentration. In contrast, at very high concentration, the MASP-2 inhibitor, which is also a poor MASP-3 inhibitor, slowed down the activation. When recombinant MASPs were added to plasma, only MASP-3 could reduce the half-life of pro-FD. Combining our quantitative data, MASP-1 and MASP-2 can be ruled out as direct pro-FD activators in resting blood; however, active MASP-3 is a very likely physiological activator.

Original languageEnglish
Pages (from-to)857-865
Number of pages9
JournalJournal of Immunology
Volume196
Issue number2
DOIs
Publication statusPublished - Jan 15 2016

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Complement Factor D
Mannose-Binding Lectin
Serine Proteinase Inhibitors
Serine Proteases
Mannose-Binding Protein-Associated Serine Proteases
DegP protease
Serum
Enzyme Precursors
Enzymes

ASJC Scopus subject areas

  • Immunology

Cite this

MASP-1 and MASP-2 do not activate pro-factor D in resting human blood, whereas MASP-3 is a potential activator : Kinetic analysis involving specific MASP-1 and MASP-2 inhibitors. / Oroszlán, Gábor; Kortvely, Elod; Szakács, Dávid; Kocsis, Andrea; Dammeier, Sascha; Zeck, Anne; Ueffing, Marius; Závodszky, P.; Pál, G.; Gál, P.; Dobó, J.

In: Journal of Immunology, Vol. 196, No. 2, 15.01.2016, p. 857-865.

Research output: Contribution to journalArticle

Oroszlán, Gábor ; Kortvely, Elod ; Szakács, Dávid ; Kocsis, Andrea ; Dammeier, Sascha ; Zeck, Anne ; Ueffing, Marius ; Závodszky, P. ; Pál, G. ; Gál, P. ; Dobó, J. / MASP-1 and MASP-2 do not activate pro-factor D in resting human blood, whereas MASP-3 is a potential activator : Kinetic analysis involving specific MASP-1 and MASP-2 inhibitors. In: Journal of Immunology. 2016 ; Vol. 196, No. 2. pp. 857-865.
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