A novel aspartic protease inhibitor inhibits the enzymatic browning of potatoes

Authors: DONG, TIANTIAN - Shandong Agricultural University; YU, CAO - Shandong Agricultural University; LI, GUANGCUN - Shandong Agricultural University; ZHU, ZIRU - Shandong Agricultural University; ZHANG, SONG - Shandong Agricultural University; Jiang, Cai-Zhong; WANG, QINGGUQ - Shandong Agricultural University

Submitted to: Postharvest Biology and Technology
Publication Type: Peer Reviewed Journal
Publication Acceptance Date: 9/8/2020
Publication Date: 2/1/2021
Citation: Dong, T., Yu, C., Li, G., Zhu, Z., Zhang, S., Jiang, C., Wang, Q. 2021. A novel aspartic protease inhibitor inhibits the enzymatic browning of potatoes. Postharvest Biology and Technology. 172. Article 111353. https://doi.org/10.1016/j.postharvbio.2020.111353.
DOI: https://doi.org/10.1016/j.postharvbio.2020.111353

Interpretive Summary: Potato (Solanum tuberosum L.) is an economically important staple crop. In recent decades, fresh-cut products including processed potato have been increasing in popularity. However, processed potato products are prone to enzymatic browning, causing a considerable loss in quality and value. Although many technologies to inhibit browning have been investigated in the processed produce industry, it is still a challenge to better solve this problem. Enzymatic browning is mainly caused by the oxidation of phenolic substrates under the catalysis of polyphenol oxidase (PPO). However, there are abundant of free amino acids (FAAs) and soluble proteins in potato tubers, which could combine with o-quinone to form brown quinones or colorless substance, thus promoting or inhibiting browning. Furthermore, proteases are also associated with enzymatic browning of potatoes. Proteases can be activated by various stresses, such as low temper- ature and physical injury, leading to the hydrolysis of endogenous proteins and the increase of free amino acids. Therefore, regulating proteases might be an important way to control enzymatic browning. Protease inhibitors, which are abundant in potato tubers and plant seeds, act as safety valves to regulate activities of proteases. However, there is little information about the effects of protease inhibitors on enzymatic browning of potatoes. In our previous studies, we found potatoes from a cultivar “Kexin #4” [bred from hybridization of ‘Pulsatilla’ (female) and ‘Katahdin’ (male)] are much more prone to browning than those from the cultivar “Kexin #13” (self-bred progeny of ‘Mira’). And a technical treatment, postharvest curing treatment (PCT), effectively inhibited potato browning after fresh-cut, though PCT increased PPO activity and phenolic com- pounds. To study the molecular mechanism underlying potatoes with different browning-susceptibilities, RNA-seq analysis was performed using these materials. Through transcriptomic analysis, the differentially expressed genes and the enriched differential terms were found mainly involved with protease inhibitors. Among these protease inhibitors, a novel aspartic protease inhibitor gene, named as StASPI, was identified in this study. Overexpression of StASPI effectively reduced enzymatic browning after cutting, significantly decreased protease activity, and reduced the accumulation of total FAAs in potatoes. Browning degrees of transgenic potato mash were increased by supplementing exogenous FAAs, but the degrees were still significantly lower than that of wild-type (WT) mash. Furthermore, overexpression of StASPI decreased PPO activity, enhanced the activities of antioxidant enzymes SOD and CAT, and reduced H2O2 and O2'– contents. These results indicated that overexpression of StASPI inhibited the enzymatic browning of potatoes, decreasing FAAs accumulation, reducing PPO activity, and enhancing activity of antioxidant enzymes. This study provides a new perspective on the strategies for inhibiting enzymatic browning of potatoes.

Technical Abstract: Enzymatic browning greatly affects the quality of fresh-cut potatoes (Solanum tuberosum L.). The transcript level of a novel aspartic protease inhibitor gene, StASPI, was found significantly higher in browning-less potatoes than in browning-susceptible potatoes, and exogenous aspartic protease inhibitor Pepstatin A inhibited the browning of potato pulp. Therefore, we speculated that StASPI plays important roles in browning-resistance of potatoes. The effects of overexpressed-StASPI on browning, total free amino acids (FAAs), antioxidant enzyme activity and ROS accumulation in potatoes were investigated in this study. Results showed that overexpression of StASPI effectively reduced enzymatic browning after cutting, significantly decreased protease activity, and reduced the accumulation of total FAAs in potatoes. Browning degrees of transgenic potato mash were increased by supplementing exogenous FAAs, but the degrees were still significantly lower than that of wild-type (WT) mash. Furthermore, overexpression of StASPI decreased PPO activity, enhanced the activities of antioxidant enzymes SOD and CAT, and reduced H2O2 and O2'– contents. These results indicated that overexpression of StASPI inhibited the enzymatic browning of potatoes, decreasing FAAs accumulation, reducing PPO activity, and enhancing activity of antioxidant enzymes. This study provides a new perspective on the strategies for inhibiting enzymatic browning of potatoes.