ABSTRACIFour aminopeptidases (APs) were separated using native polyacrylamide gel electrophoresis of cell-free extracts and the stromal fractions of isolated chloroplasts prepared from primary barley (Hordeum vulgare L., var Numar) leaves. Activities were identified using a series of aminoacyl-,B-naphthylamide derivatives as substrates. AP1, 2, and 3 were found in the stromal fraction of isolated chloroplasts with respective molecular masses of 66.7, 56.5, and 54.6 kilodaltons. AP4 was found only in the cytoplasmic fraction. No AP activity was found in vacuoles of these leaves. It was found that 50% of the L-Leu-#-naphthylamide and 25% of the L-Arg-ji-naphthylamide activities were localized in the chloroplasts. Several AP activities were associated with the membranes of the thylakoid fraction of isolated chloroplasts. API, 2, and 4 reacted against a broad range of substrates, whereas AP3 hydrolyzed only LArg-B-naphthylamide. Only AP2 hydrolyzed L-Val-a-naphthylamide. Since AP2 and AP3 were the only ones reacting against Val-#-naphthylamide and Arg-8-naphthylamide, respectively, several protease inhibitors were tested against these substrates using a stromal fraction from isolated chloroplasts as the source of the two APs. Both APs were sensitive to both metallo and sulfhydryl type inhibitors. Although AP activity decreased as leaves senesced, no new APs appeared on gels during senescence and none disappeared.Soluble proteins are rapidly degraded or mobilized in the early phases of leaf senescence. Chloroplast proteins are among the first to be degraded, and chloroplast constituents disappear faster than the chloroplast organelles (10,13,28). This suggests that chloroplasts contain proteolytic activities capable of degrading their protein constituents. Such proteolytic activities in chloroplasts have, in fact, been found. Isolated chloroplasts from barley (3) and soybean (19) were capable of degrading RuBPCase4; an ATP-dependent proteolytic activity was located in thylakoids from pea chloroplasts (8, 11), and one endoprotease and three APs were found in the stromal fraction of pea chloroplasts (9); endoproteolytic activity against RuBPCase was found in the stromal fractions of chloroplasts from barley leaves (22,23) Nettleton et al. (17) and Tang and Huffaker (23) found proteolytic activities associated with the thylakoid fractions of chloroplasts from wheat and barley leaves, respectively, capable of degrading RuBPCase. In wheat, 50% of the AP activity was localized in chloroplasts with the remainder in the cytoplasm (30).To better understand the degradation ofchloroplastic proteins, it is necessary to identify and characterize the proteases and their subcellular localization. Recent information shows that the in vivo half-life of some bacterial proteins is a function of their amino terminal amino acid residue (2). If future work shows this applicable to plants, APs may be important in the degradation of chloroplast protein. In this paper, we have identified four APs from barley leaves that are localized in both...

Erythromycin (Ery) is a commonly used veterinary drug that prevents infections and promotes the growth of farm animals. Ery is often detected in agricultural fields due to the effects of manure application in the ecosystem. However, there is a lack of information on Ery toxicity in crops. In this study, we performed a comparative proteomic analysis to identify the molecular mechanisms of Ery toxicity during seedling growth based on our observation of a decrease in chlorophyll (Chl) contents using Brassica campestris. A total of 452 differentially abundant proteins (DAPs) were identified including a ribulose-1,5-bisphosphate carboxylase (RuBisCO). The proteomic analysis according to gene ontology (GO) classification revealed that many of these DAPs responding to Ery treatment functioned in a cellular process and a metabolic process. The molecular function analysis showed that DAPs classified within catalytic activity were predominantly changed by Ery, including metabolite interconversion enzyme and protein modifying enzyme. An analysis of functional pathways using MapMan revealed that many photosynthesis components were downregulated, whereas many protein biosynthesis components were upregulated. A good relationship was observed between protein and transcript abundance in a photosynthetic pathway, as determined by qPCR analysis. These combined results suggest that Ery affects plant physiological activity by downregulating protein abundance in the photosynthetic pathway.

Saponarin content in barley sprouts may vary greatly with environmental conditions, such as climate, leading to difficulty in uniformly producing saponarin-rich barley sprouts in situ farmlands throughout the year. This research was an early attempt to identify the optimal conditions of various climatic factors, such as temperature, light, and humidity according to seasonal change, for maximizing the saponarin content of sprouted barley through the two-year field experiment. As a result, the growth index, as leaf length relative to growth period, of barley sprouts varied greatly with sowing time, and they tended to decrease with an increase in the ambient temperature, such as average daily temperature. In contrast, higher saponarin contents were observed in the sprouts collected in March, April, September, and October than those collected from May to August. We also found significantly positive correlations of saponarin content with daily temperature range and average light period, indicating that they could be decisive climatic factors for the production of barley sprouts with a higher saponarin content. Interestingly, the polynomial relationship between saponarin yield and leaf length showed the highest yield with 2.18 mg plant−1 at 15.9 cm in length, suggesting a best cutting time for the production of saponarin-rich barely sprouts based on the leaf length. Overall, the decisive climatic factors according to seasonal change for saponarin biosynthesis may be considered to be daily temperature differences and light hours.

Veterinary antibiotics such as sulfonamides are widely used to increase feed efficiency and to protect against disease in livestock production. The sulfonamide antimicrobial mechanism involves the blocking of folate biosynthesis by inhibiting bacterial dihydropteroate synthase (DHPS) activity competitively. Interestingly, most treatment antibiotics can be released into the environment via manure and result in significant diffuse pollution in the environment. However, the physiological effects of sulfonamide during plant growth and development remain elusive because the plant response is dependent on folate biosynthesis and the concentration of antibiotics. Here, we present a chemical interaction docking model between Napa cabbage (Brassica campestris) DHPS and sulfamethoxazole and sulfamethazine, which are the most abundant sulfonamides detected in the environment. Furthermore, seedling growth inhibition was observed in lentil bean (Lens culinaris), rice (Oryza sativa), and Napa cabbage plants upon sulfonamide exposure. The results revealed that sulfonamide antibiotics target plant DHPS in a module similar to bacterial DHPS and affect early growth and the development of crop seedlings. Taking these results together, we suggest that sulfonamides act as pollutants in crop fields.

Saponarin (SA) is a major di-C-glycosyl-O-glycosyl flavone, which is predominantly accumulated in the young green leaves of barley (Hordeum vulgare L.), with numerous biological functions in plants, such as protection against environmental stresses. Generally, SA synthesis and its localization in the mesophyll vacuole or leaf epidermis are largely stimulated in response to biotic and abiotic stresses to participate in a plant’s defense response. In addition, SA is also credited for its pharmacological properties, such as the regulation of signaling pathways associated with antioxidant and anti-inflammatory responses. In recent years, many researchers have shown the potential of SA to treat oxidative and inflammatory disorders, such as in protection against liver diseases, and reducing blood glucose, along with antiobesity effects. This review aims to highlight natural variations of SA in plants, biosynthesis pathway, and SA’s role in response to environmental stress and implications in various therapeutic applications. In addition, we also discuss the challenges and knowledge gaps concerning SA use and commercialization.