LeSBT1, a Subtilase from Tomato Plants

Abstract

The cDNA of a tomato subtilase designated LeSBT1 was cloned from a tomato flower cDNA library. The deduced amino acid sequence indicated for LeSBT1 the structure of a prepro-protein targeted to the secretory pathway by virtue of an amino-terminal signal peptide. LeSBT1 was expressed in the baculovirus/insect cell system and a processed 73-kDa form of LeSBT1, lacking both signal peptide and prodomain, was purified to homogeneity from culture supernatants. This 73-kDa LeSBT1, however, lacked proteolytic activity. Zymogen activation to yield 68-kDa LeSBT1 required the additional processing of an amino-terminal autoinhibitory peptide in a strictly pH-dependent manner. Mature 68-kDa LeSBT1 showed highest activity at acidic pH consistent with its presumed localization in the apoplast of the plant cell. In comparison to other plant subtilases, LeSBT1 exhibited a narrower substrate specificity in that it cleaves only polypeptide substrates preferentially but not exclusively carboxyl-terminal of glutamine residues. The possible involvement of LeSBT1 in selective proprotein processing is discussed with reference to the related mammalian proprotein convertases.

The Genome Sequence of the Tomato-Pathogenic ActinomyceteClavibacter michiganensissubsp.michiganensisNCPPB382 Reveals a Large Island Involved in Pathogenicity

“…sepedonicus only two homologs are present (CMS0598 and CMS0597). The encoded proteins are interesting as they have high similarity to the SBT1, SBT2, and P69 subtilases of tomato (28,29). The exact functions of these tomato proteases are unknown, but some of them have been implicated in wound and pathogen responses (29).…”

Purification and Characterization of Serine Proteases That Exhibit Caspase-Like Activity and Are Associated with Programmed Cell Death in Avena sativa

“…Gene families of subtilisin-like Ser proteases have been identified in several plant species. Lycopersicon esculentum (tomato) has had 10 genes for subtilisin-like Ser proteases thus far cloned and characterized (Vera and Conejero, 1988;Tornero et al, 1997;Jordá et al, 1999Jordá et al, , 2000Meichtry et al, 1999;Janzik et al, 2000;Riggs et al, 2001). Whereas Arabidopsis (Arabidopsis thaliana) has had only four genes characterized (Neuteboom et al, 1999;Berger and Altmann, 2000;Tanaka et al, 2001;Hamilton et al, 2003), at least 10 more genes have been annotated as being subtilisin-like Ser proteases.…”

“…Autoproteolysis by SAS-1 and SAS-2 is not unique among plant subtilisin-like Ser proteases because two other proteases have been shown to undergo a form of self-catalyzed proteolysis. LeSBT1, a 73-kD protease from L. esculentum, is autoprocessed into a 68-kD protease at pH 5.0 and 6, whereas it completely autodegrades at pH 4.0 (Janzik et al, 2000), and hordolisin from Hordeum vulgare (barley) was autoprocessed from 74 to 62 kD during storage at ÿ208C (Terp et al, 2000).…”

“…RFU, relative fluorescence units. and Distribution1 (SDD1) (Berger and Altmann, 2000), Tomato subtilase1 (LeSBT1) (Janzik et al, 2000), 50-kD Systemin Binding Protein (Schaller and Ryan, 1994), and Ara12 (Hamilton et al, 2003) are described as possessing greater substrate specificity and are thought to function as processive proteases. Until more is known about the biochemical and physiological properties of plant subtilisin-like Ser proteases, the biological significance of the difference between a degradative or processive function may prove challenging to determine.…”

Systemic wound signaling in plants: A new perception

“…Systemin would generate more JA, resulting in the amplification of systemin and oxylipins as a cascade along the stems and petioles. The process would eventually be limited by the presence of extracellular systemininactivating enzymes (22). A signal cascade initiated in a wounded wild-type rootstock grafted to an spr-2 scion would amplify all of the signals through the vascular bundles to the graft junction but could not proceed along the spr-2 stem.…”