Calcium requirements of residual protease in Bacillus subtilis DB104

Longo, Alessandra; Snay, J.; Zimmer, Lee A.; Ataai, Mohammad M.

doi:10.1007/bf01024719

Cited by 1 publication

(2 citation statements)

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“…Production of the enzyme was performed in B. subtilis strain DB104 which lacks native extracellular neutral and serine proteases. [16] Peptiligase is translated as a precursor and needs to be processed to obtain the mature protein. It was reported that a helper protease, i.e., native subtilisin, is required for processing of subtiligase, due to its reduced protease activity.…”

Section: Resultsmentioning

confidence: 99%

“…The gene encoding peptiligase was constructed by site‐directed mutagenesis and cloned into the Escherichia coli/Bacillus subtilis shuttle vector pBS42. Production of the enzyme was performed in B. subtilis strain DB104 which lacks native extracellular neutral and serine proteases 16. Peptiligase is translated as a precursor and needs to be processed to obtain the mature protein.…”

Section: Resultsmentioning

confidence: 99%

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Peptiligase, an Enzyme for Efficient Chemoenzymatic Peptide Synthesis and Cyclization in Water

Toplak

Nuijens²,

Quaedflieg³

et al. 2016

Adv Synth Catal

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We describe a novel, organic cosolventstable and cation-independent engineered enzyme for peptide coupling reactions. The enzyme is a variant of a stable calcium-independent mutant of subtilisin BPN', with the catalytic Ser212 mutated to Cys and Pro216 converted to Ala. The enzyme, called peptiligase, catalyzes exceptionally efficient peptide coupling in water with a surprisingly high synthesis over hydrolysis (S/H) ratio. The S/H ratio of the peptide ligation reaction is correlated to the length of the peptide substrate and proved to be > 100 for the synthesis of a 13-mer peptide, which corresponds to > 99% conversion to the ligated peptide product and < 1% hydrolytic side-reaction. Furthermore, peptiligase does not require a particular recognition motif resulting in a broadly applicable and traceless peptide ligation technology. Peptiligase is very robust, easy to produce in Bacillus subtilis, and its purification is straightforward. It shows good activity and stability in the presence of organic cosolvents and chelating or denaturing agents, enabling the ligation of poorly soluble (hydrophobic) or folded peptides. This enzyme could be useful for the (industrial) synthesis of diverse (pharmaceutical) peptides. In addition, peptiligase is able to efficiently catalyze headto-tail peptide cyclization reactions.

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Section: Resultsmentioning

confidence: 99%

Section: Resultsmentioning

confidence: 99%