Reaction of nerve growth factor gamma and 7S nerve growth factor complex with human and murine alpha 2-macroglobulin.
The kallikrein-like serine proteinase nerve growth factor gamma (NGF-gamma) reacted with the plasma proteinase inhibitor human alpha 2-macroglobulin (h alpha 2M). The h alpha 2M subunits were cleaved, the electrophoretic mobility of h alpha 2M in nondenaturing polyacrylamide gels was increased, and the intrinsic fluorescence of h alpha 2M was increased with a slight blue-shift. These changes are well-characterized components of the alpha 2M/proteinase reaction mechanism. In N alpha-benzoyl-DL-arginine p-nitroanilide (BAPNA) hydrolysis experiments, the catalytic efficiency (kcat/KM) of the h alpha 2M-NGF-gamma complex was decreased by 98.5% compared with free NGF-gamma. This decrease is unique since other alpha 2M-proteinase complexes retain significant amidase activity. For comparison, we determined that the catalytic efficiency of alpha 2M-trypsin is decreased by 58% compared with free trypsin under equivalent conditions. The rate of NGF-gamma inhibition by h alpha 2M was (1.0 +/- 0.1) x 10(4) M-1 s-1 as determined by BAPNA hydrolysis. A similar value was determined by monitoring the change in intrinsic fluorescence. NGF-gamma, which was bound within the intact 7S NGF complex, also reacted with h alpha 2M, albeit at a very slow rate. This reaction may have depended exclusively on slow reversible dissociation of NGF-gamma from the 7S complex. NGF-gamma was rapidly inhibited by murine alpha 2M (m alpha 2M).(ABSTRACT TRUNCATED AT 250 WORDS)
Wolf
B B
BB
Department of Pathology, University of Virginia Health Sciences Center, Charlottesville 22908.
Vasudevan
J
J
Gonias
S L
SL
D000818:Animals; D001586:Benzoylarginine Nitroanilide / Q000378:metabolism; D002863:Chromogenic Compounds / Q000378:metabolism; D004591:Electrophoresis, Polyacrylamide Gel; D006801:Humans; D006868:Hydrolysis; D007700:Kinetics; D051379:Mice; D009414:Nerve Growth Factors / Q000737:chemistry / Q000378:metabolism*; D011487:Protein Conformation; D012697:Serine Endopeptidases / Q000737:chemistry / Q000378:metabolism*; D013050:Spectrometry, Fluorescence; D014357:Trypsin / Q000378:metabolism; D000511:alpha-Macroglobulins / Q000378:metabolism* / Q000494:pharmacology
C013614:7S nerve growth factor complex; C488871:7S nerve growth factor protein, human; D002863:Chromogenic Compounds; D009414:Nerve Growth Factors; D000511:alpha-Macroglobulins; D001586:Benzoylarginine Nitroanilide; D012697:Serine Endopeptidases; C079910:nerve growth factor, gamma; D014357:Trypsin