Fig. 3: Heterologous expression of CsKPI in E. coli for determination of enzymatic properties and subcellular localization. | Horticulture Research

Fig. 3: Heterologous expression of CsKPI in E. coli for determination of enzymatic properties and subcellular localization.

From: Duplication and transcriptional divergence of three Kunitz protease inhibitor genes that modulate insect and pathogen defenses in tea plant (Camellia sinensis)

Fig. 3

a SDS-PAGE image of the purified CsKPI1, CsKPI2, and CsKPI3 recombinant proteins: 1, CsKPI1; 2, CsKPI2; and 3, CsKPI3. The solid box shows the expected protein positions. b Representative HPLC analyses of enzymatic reaction products after incubating recombinant CsKPI proteins with trypsin and N-α-benzoyl-dl-arginine-4-nitroanilide (BAPNA). Traces 3–6 represent the products from reactions with the recombinant proteins GST, CsKPI1, CsKPI2, and CsKPI3, respectively; traces 1 and 2 represent a standard (PNA) and a blank control group, respectively. c Quantitative determination of the PNA content. Bars indicate the means ± SD (n = 3) of three biological replicates. Asterisks indicate the significance level (*P < 0.05, **P < 0.01) based on Tukey’s honestly significant difference test. d Subcellular localization of three CsKPIs. Rows 1–4 show confocal images for the 35:GFP, 35:CsKPI1-GFP, 35:CsKPI2-GFP, and 35:CsKPI3-GFP constructs, which are each transiently expressed in Arabidopsis protoplasts. Scale bar = 10 μm

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