PLANT PHYSIOLOGY , Vol 106, Issue 4 1461-1469, Copyright © 1994 by American Society of Plant Biologists
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METABOLISM AND ENZYMOLOGY |
Tomato Fruit Polygalacturonase Isozyme 1 (Characterization of the [beta] Subunit and Its State of Assembly in Vivo)
T. Moore and A. B. Bennett
Mann Laboratory, Department of Vegetable Crops, University of California, Davis, California 95616
Polygalacturonase isozyme 1 (PG1) is a heterodimer comprising a catalytic
and noncatalytic or [beta] subunit, whereas polygalacturonase isozyme 2
(PG2) comprises only the catalytic subunit. To assess the state of assembly
of PG1 in vivo, both subunits were purified to homogeneity and used to
study assembly of the heterodimer. PG1 could be reconstituted in vitro from
purified [beta] subunit and purified PG2 under a wide range of salt and pH
conditions, and PG1 reconstituted in vitro was indistinguishable from PG1
isolated from tomato (Lycopersicon esculentum) fruit. Specific antibodies
indicated that the [beta] subunit was present in fruit of all developmental
stages, but absent in vegetative tissue. The state of assembly of PG1 in
vivo was tested based on the differential thermal stability of PG1 and PG2
by heating segments of ripe fruit pericarp tissue. Temperatures well below
those required to inactivate PG1 in vitro caused the loss of activity of
both PG1 and PG2, suggesting that only heat-labile PG2 is present in vivo.
In addition, when extracts of ripe fruit were rigorously maintained and
analyzed at 4[deg]C, PG1 was absent or barely detectable. These results are
consistent with the hypothesis that PG1 can assemble spontaneously and is
essentially absent in intact tomato fruit but forms artifactually from PG2
and the [beta] subunit during the extraction of tomato fruit tissue when
low temperatures are not rigorously maintained.
