Structural elements that modulate the substrate specificity of plant purple acid phosphatases: Avenues for improved phosphorus acquisition in crops

Abstract

Experimental details for the ITC assay. The enthalpy change (ΔH) associated with the reaction of rkbPAP with p-NPP was determined by injecting 10 µL of p-NPP (20 mM) into the reaction cell (200 µL) containing 0.95 µM of rkbPAP, and by allowing the reaction to proceed to completion.The reaction is exothermic as indicated by the negative value of the heat pulse (dQ/dT); the value of ΔH can be calculated by integrating the area under the curve in Fig. S2a (minus the endothermic heat associated with the dilution) divided by the amount of p-NPP hydrolyzed in the cell. The value for the endothermic heat generated by the dilution was obtained by injecting 10 µL of p-NPP (20 mM) into the reaction cell containing only buffer: ΔH = 3089 ± 52 µcal/µmol (i.e. 12.9 ± 0.22 kJ/mol), in good agreement with the corresponding ΔH value determined by enzymatic assays (13.2 ± 0.18 kJ/mol) [1]. In this study, we focused on measuring initial catalytic rates in order to minimize the effect of the reaction product and competitive inhibitor phosphate [2][3][4][5][6].Different concentrations of p-NPP were prepared and 8 µL of 0.24 µM rkbPAP were injected into the substrate solutions. Fig. S2b shows a typical calorimetric trace of a reaction. The initial rates determined in µcal/s, when divided by ΔH (µcal/µmol), are transformed into µmol/s, representing the amount of p-NPP hydrolyzed per second in the cell. These rates were subsequently plotted against the p-NPP concentration and fitted to the Michaelis-Menten equation (Eq. 1; Fig. S2c), resulting in a kcat value (i.e. Vmax/[PAP]) of ~190 s -1 and a Michaelis constant Km of ~4 mM.

Potato root and leaf phosphatase activity in response to P deprivation

“…Recently, Feder et al. (2020) identified elements in the protein structure of purple acid phosphatases that determine substrate preferences and therefore biological functions. Only a few phosphatases are secreted by roots as known from rice, soybean, poplar, and Arabidopsis (Kavka et al., 2021; Lu et al., 2016; Robinson, Park et al., 2012; Tran, Qian et al., 2010; Wang et al., 2014; Zhu et al., 2020).…”

Genome-Wide Characterization and Evolutionary Analyses of Purple Acid Phosphatase (PAP) Gene Family with Their Expression Profiles in Response to Low Phosphorus Stresses in Moso Bamboo (Phyllostachys edulis)

“…The hydrolytic activity is called nonspecific PAP. It plays an important role in maintaining phosphorus homeostasis by hydrolyzing various organophosphorus compounds under weak acid conditions [4,52]. This inspired us to analyze the PAP gene family and their expression patterns of Moso bamboo, hoping to provide references for revealing the low-phosphorus adaptation mechanism of Moso bamboo and provide key candidate genes for the genetic engineering supported breeding of Moso bamboo.…”

“…However, the demand for nutrients for the rapid growth of Moso bamboo is very high, and the sufficient supply of soil nutrients, especially phosphorus (P), limits the growth of these plants. Phosphorus is an essential mineral element for plant growth and development and plays an important role in energy transfer [4], Forests 2021, 12, 2 of 19 signal transduction [5], photosynthesis and respiration [6]. Plants mainly absorb inorganic soluble phosphate in the soil through the root system but cannot directly absorb a large amount of organic phosphorus, such as phosphate ester and phosphate anhydride [7].…”

Phosphorus Mobilizing Capacity of Selected Grain Legumes Grown Under Phosphorus-Deficient Conditions

“…Replacement of membrane phospholipids by lipids that do not contain P, could save P for other cellular processes when plants are grown under Pdeficiency. Moreover, vacuolar purple acid phosphatases scavenge P from intracellular Pcontaining compounds during prolong Pdeficiency (Feder et al, 2020;Luo et al, 2020). Some of these mechanisms might have come into effect in determining the enhanced use efficiency of acquired P by black gram.…”