Characterisation of an alpha-galactosidase with potential relevance to ripening related texture changes.
alpha-Galactosidase (EC 3.2.1.22) from ripe papaya (Carica papaya L.) fruit was fractionated by a combination of ion exchange and gel filtration chromatography into three forms, viz., alpha-galactosidase 1, 2 and 3. The predominant isoform, alpha-gal 2, was probably a tetramer with a native molecular mass of about 170 kDa and 52 kDa-sized subunits and an estimated pI of 7.3. The subunit's N-terminal amino acid sequence shared high identity (97%) with the deduced sequence of a papaya cDNA clone encoding a putative alpha-galactosidase PAG2 as well as with an Ajuga reptans L. GGT1 clone encoding a galactan: galactan galactosyltransferase (66%). During ripening, alpha-galactosidase activity increased concomitantly with firmness loss and this increase was largely ascribed to alpha-gal 2. The protein level of alpha-gal 2 as estimated by immunoblot was low in developing fruits and generally increased with ripening. alpha-Galactosidase 2 also had the ability to markedly catalyse increased pectin solubility and depolymerisation while the polymers were still structurally attached to the cell walls mimicking, in part, the changes that occur during ripening. The close correlation between texture changes, alpha-gal 2 activity and protein levels as well as capability to modify intact cell walls suggest that the enzyme might contribute to papaya fruit softening during ripening. The purported mechanism of alpha-gal 2 action as a softening enzyme was discussed in terms of its functional capacity as a glycanase or perhaps, as a transglycosylase.
Soh
Chin-Pin
CP
School of Biosciences and Biotechnology, Faculty of Science and Technology, Universiti Kebangsaan Malaysia, 43600 Bangi, Selangor, Malaysia.
Ali
Zainon Mohd
ZM
Lazan
Hamid
H
D000595:Amino Acid Sequence; D029441:Carica / Q000737:chemistry / Q000201:enzymology* / Q000254:growth & development; D002412:Cations / Q000378:metabolism; D002473:Cell Wall / Q000201:enzymology; D005638:Fruit / Q000201:enzymology; D007527:Isoenzymes / Q000737:chemistry / Q000302:isolation & purification; D007700:Kinetics; D008969:Molecular Sequence Data; D009005:Monosaccharides / Q000378:metabolism; D009598:Nitrophenylgalactosides / Q000378:metabolism; D010368:Pectins / Q000737:chemistry; D016415:Sequence Alignment; D017386:Sequence Homology, Amino Acid; D013379:Substrate Specificity; D000519:alpha-Galactosidase / Q000737:chemistry* / Q000302:isolation & purification / Q000502:physiology
D002412:Cations; D007527:Isoenzymes; D009005:Monosaccharides; D009598:Nitrophenylgalactosides; C059200:4-nitrophenylgalactoside; D010368:Pectins; D000519:alpha-Galactosidase