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Purification and properties of an aspartic protease from potato tuber that is inhibited by a basic chitinase
Author(s) -
Guevara María G.,
Oliva Claudia R.,
Machinandiarena Milagros,
Daleo Gustavo R.
Publication year - 1999
Publication title -
physiologia plantarum
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.351
H-Index - 146
eISSN - 1399-3054
pISSN - 0031-9317
DOI - 10.1034/j.1399-3054.1999.106203.x
Subject(s) - pepstatin , biochemistry , ammonium sulfate precipitation , proteolysis , protease , chitinase , chromatography , affinity chromatography , aspartic acid , polyacrylamide gel electrophoresis , size exclusion chromatography , gel electrophoresis , chemistry , biology , enzyme , amino acid
A protease was isolated from potato ( Solanum tuberosum L. cv. Huinkul) tuber disks after 24 h of aeration when proteolysis is markedly increased. Purification was performed by ammonium sulfate precipitation, ion exchange chromatography, and affinity chromatography. A size of 40 kDa was estimated by sodium dodecyl sulphate‐polyacrylamide gel electrophoresis (SDS‐PAGE) and gel filtration, it is monomeric and its properties are consistent with those of aspartic proteinases (EC 3.4.23): it had a pH optimum between 4 and 5 and it was inhibited by pepstatin. Partial homology with other plant aspartic proteinases was observed in two sequenced tryptic fragments. It binds to Sepharose‐concanavalin A and can be eluted with α ‐methyl mannoside, indicating that it is possibly glycosylated. Unlike other aspartic proteinases from Solanaceae that degrade pathogenesis‐related proteins, it is unable to cleave a basic chitinase from potato. Moreover, this aspartic protease is strongly inhibited by the basic chitinase; the 50% inhibition is obtained when the molar ratio approaches 1, the same as with pepstatin. The interaction between this aspartic protease and a new type of endogenous inhibitor may be an interesting starting point to study the regulation of these aspartic proteases during stress.