Structural basis of cucumisin protease activity regulation by its propeptide

Sotokawauchi, Ami; Kato-Murayama, M.; Murayama, Kazutaka; Hosaka, Toshiaki; Maeda, Isamu; Onjo, Michio; Ohsawa, Noboru; Kato, Dai Ichiro; Arima, Kazunari; Shirouzu, Mikako

doi:10.1093/jb/mvw053

Cited by 12 publications

(10 citation statements)

References 39 publications

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“…3C). The data suggest that the neo-C terminus of SPI-1 remains bound and blocks the active site of the protease in a product-like manner as reported for PPs of bacterial and plant SBTs (20,22) and inferred for fungal I9s (28).…”

Section: Characterization Of the Spi-1⅐sbt413 Complexsupporting

confidence: 61%

“…Sequence variability between PPs and I9s was greatest at their N and C termini. As judged from the crystal structure of the cucumisin⅐prodomain complex, the N terminus of the PP does not seem to be important for binding (22). Therefore, it may not be subject to evolutionary selection, possibly explaining N-terminal variability in length and sequence.…”

Section: Figure 1 Phylogenetic Relationship Of Propeptides and I9 Inmentioning

confidence: 98%

“…Crystal structure analysis of PP⅐SBT complexes revealed a common mechanism of autoinhibition for bacterial subtilisins (8,20), mammalian PCSK9 (21), and cucumisin, an abundant SBT in melon fruits (22). In each case, the ␤-sheet of the propeptide packs against two parallel surface helices of the respective protease, whereas its C terminus binds to the nonprime subsites of the catalytic center in a product-like manner (subsites up-and downstream of the cleavage site are called nonprime and prime subsites, respectively, according to Schechter and Berger (65)).…”

mentioning

confidence: 99%

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A novel subtilase inhibitor in plants shows structural and functional similarities to protease propeptides

Hohl

Stintzi

Schaller

2017

Journal of Biological Chemistry

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The propeptides of subtilisin-like serine proteinases (subtilases, SBTs) serve dual functions as intramolecular chaperones that are required for enzyme folding and as inhibitors of the mature proteases. SBT propeptides are homologous to the I9 family of protease inhibitors that have only been described in fungi. Here we report the identification and characterization of subtilisin propeptide-like inhibitor 1 (SPI-1) from Sequence similarity and the shared β-α-β-β-α-β core structure identified SPI-1 as a member of the I9 inhibitor family and as the first independent I9 inhibitor in higher eukaryotes. SPI-1 was characterized as a high-affinity, tight-binding inhibitor of subtilase SBT4.13 with and values in the picomolar range. SPI-1 acted as a stable inhibitor of SBT4.13 over the physiologically relevant range of pH, and its inhibitory profile included many other SBTs from plants but not bovine chymotrypsin or bacterial subtilisin A. Upon binding to SBT4.13, the C-terminal extension of SPI-1 was proteolytically cleaved. The last four amino acids at the newly formed C terminus of SPI-1 matched both the cleavage specificity of SBT4.13 and the consensus sequence of SBTs at the junction of the propeptide with the catalytic domain. The data suggest that the C terminus of SPI-1 acts as a competitive inhibitor of target proteases as it remains bound to the active site in a product-like manner. SPI-1 thus resembles SBT propeptides with respect to its mode of protease inhibition. However, in contrast to SBT propeptides, SPI-1 could not substitute as a folding assistant for SBT4.13.

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Section: Characterization Of the Spi-1⅐sbt413 Complexsupporting

confidence: 61%

Section: Figure 1 Phylogenetic Relationship Of Propeptides and I9 Inmentioning

confidence: 98%

mentioning

confidence: 99%

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A novel subtilase inhibitor in plants shows structural and functional similarities to protease propeptides

Hohl

Stintzi

Schaller

2017

Journal of Biological Chemistry

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“…Additional exosite interactions of the prodomain with two parallel surface helices of the protease contribute to complex formation ( Fig. 1; Sotokawauchi et al, 2017). In contrast to SlSBT3, which shows high specificity for its own prodomain (Meyer et al, 2016b), cucumisin is also inhibited by distantly related prodomains of SBTs from rice and Arabidopsis (Nakagawa et al, 2010).…”

Section: Biochemistry and Structure Of Plant Sbtsmentioning

confidence: 99%

From structure to function – a family portrait of plant subtilases

et al. 2017

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Contents Summary901I.Introduction901II.Biochemistry and structure of plant SBTs902III.Phylogeny of plant SBTs and family organization903IV.Physiological roles of plant SBTs905V.Conclusions and outlook911Acknowledgements912References912 Summary Subtilases (SBTs) are serine peptidases that are found in all three domains of life. As compared with homologs in other Eucarya, plant SBTs are more closely related to archaeal and bacterial SBTs, with which they share many biochemical and structural features. However, in the course of evolution, functional diversification led to the acquisition of novel, plant‐specific functions, resulting in the present‐day complexity of the plant SBT family. SBTs are much more numerous in plants than in any other organism, and include enzymes involved in general proteolysis as well as highly specific processing proteases. Most SBTs are targeted to the cell wall, where they contribute to the control of growth and development by regulating the properties of the cell wall and the activity of extracellular signaling molecules. Plant SBTs affect all stages of the life cycle as they contribute to embryogenesis, seed development and germination, cuticle formation and epidermal patterning, vascular development, programmed cell death, organ abscission, senescence, and plant responses to their biotic and abiotic environments. In this article we provide a comprehensive picture of SBT structure and function in plants.

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“…The tertiary structures of subtilases have been reported earlier, i.e. the cucumisin from melon fruits (Protein Data Bank (PDB) code 3VTA and 4YN3) 30,31 , and SlSBT3 from tomato (PDB code 3I6S) 32 . However, the protein structural information of SBT2 (the earliest-diverging subtilase of embryophytes; Fig.…”

Section: Resultsmentioning

confidence: 82%

Molecular evidence for origin, diversification and ancient gene duplication of plant subtilases (SBTs)

Wang

et al. 2019

Sci Rep

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Plant subtilases (SBTs) are a widely distributed family of serine proteases which participates in plant developmental processes and immune responses. Although SBTs are divided into seven subgroups in plants, their origin and evolution, particularly in green algae remain elusive. Here, we present a comprehensive large-scale evolutionary analysis of all subtilases. The plant subtilases SBT1-5 were found to be monophyletic, nested within a larger radiation of bacteria suggesting that they originated from bacteria by a single horizontal gene transfer (HGT) event. A group of bacterial subtilases comprising representatives from four phyla was identified as a sister group to SBT1-5. The phylogenetic analyses, based on evaluation of novel streptophyte algal genomes, suggested that the recipient of the HGT of bacterial subtilases was the common ancestor of Coleochaetophyceae, Zygnematophyceae and embryophytes. Following the HGT, the subtilase gene duplicated in the common ancestor and the two genes diversified into SBT2 and SBT1, 3–5 respectively. Comparative structural analysis of homology-modeled SBT2 proteins also showed their conservation from bacteria to embryophytes. Our study provides the first molecular evidence about the evolution of plant subtilases via HGT followed by a first gene duplication in the common ancestor of Coleochaetophyceae, Zygnematophyceae, and embryophytes, and subsequent expansion in embryophytes.

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Structural basis of cucumisin protease activity regulation by its propeptide

Cited by 12 publications

References 39 publications

A novel subtilase inhibitor in plants shows structural and functional similarities to protease propeptides

A novel subtilase inhibitor in plants shows structural and functional similarities to protease propeptides

From structure to function – a family portrait of plant subtilases

Molecular evidence for origin, diversification and ancient gene duplication of plant subtilases (SBTs)

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