[This corrects the article DOI: 10.1016/j.aninu.2019.11.003.].

Background:: Microbial phytases are added to animal feed to hydrolyze phytic acid (myoinositol hexakisphosphate, IP6) and phytate (salt of phytic acid) increasing phosphorus bioavailability. Novel phytases with enhanced bio-efficacy are being developed. Objective:: To characterize the biochemical and enzymatic properties of a novel consensus bacterial 6- phytase and its variant (PhyG), produced in Trichoderma reesei. Methods:: The in vitro specific activity, kinetic parameters, pH-activity profiles (relative to pH5.5), IP6 degradation, hydrolysis products and phosphate release of the phytases were determined using sodium phytate substrate. Melting point (Tm) was determined by differential scanning calorimetry and thermostability assessed by measuring residual activity at different temperatures. In vivo effects of PhyG supplementation at 0 to 1,000 FTU/kg on ileal IP6 digestibility and IP ester concentrations were determined in piglets. Results:: Both phytases exhibited pH optima of 3.5-4.5, high relative activity over a wide pH range (pH2.0-5.0), and substantial relative activity at pH1.5. At pH3.0, the specific activity of the PhyG variant was 1487 U/mg protein and at pH3.5 the kinetic constants were 240 μM (Km) and 1873 s-1 (Kcat). The hydrolysis of IP6 by both phytases was rapid. The major initial hydrolysis product was DLI( 1,2,3,4,5)P5, designating the phytases as bacterial 6-phytases (EC 3.1.3.26). Hydrolysis occurred at the D-3 (L-1) position in ~30% of instances, indicating a dual-specificity. Conclusion: Both phytases showed high thermostability compared to wild type and existing commercial bacterial 6-phytases; PhyG exhibited 95% residual activity after 20 min incubation at 85.4ºC (pH5.5), Tm50 of ~93.2ºC and Tm of 98.8ºC. In vivo, PhyG at 1,000 FTU/kg achieved an ileal digestibility of IP6 of 89.3%.

The utility of a next generation biosynthetic bacterial 6-phytase (PhyG) in restoring bone ash, bone phosphorus (P) content and performance in piglets depleted in P was evaluated. A total of 9 treatments were tested as follows. Treatment 1, a negative control (NC) diet; treatments 2, 3, 4, NC supplemented with 250, 500 or 1,000 FTU/kg of PhyG; treatments 5, 6, NC supplemented with 500 or 1,000 FTU/kg of a commercial Buttiauxella sp phytase (PhyB); treatments 7, 8, 9, NC supplemented with monocalcium phosphate (MCP) to provide 0.7, 1.4 and 1.8 g/kg digestible P, equating to a digestible P content of 1.8, 2.5 and 2.9 g/kg. The latter constituting the positive control (PC) diet with adequate P and calcium (Ca). The NC was formulated without inorganic P (1.1 g digestible P/kg) and reduced in Ca (5.0 g/kg). Additional limestone was added to treatments 7 to 9 to maintain Ca-to-P ratio between 1.2 and 1.3. A total of 162 crossed Pietrain × (Large White × Landrace) 21-d-old piglets (50% males and 50% females) were fed adaptation diets until 42 d old and then assigned to pens with 2 pigs/pen and 9 pens/treatment in a completely randomized block design. Piglets were fed mash diets based on corn and soybean meal ad libitum for 28 d. At the end of the study, one piglet perpen was euthanized and the right feet collected for determination of bone strength, bone ash and mineral content. Compared with the PC, the NC group had reduced average daily gain (ADG) and increased feed conversion ratio (FCR) during all growth phases and overall, and at d 28 (70 d old) NC pigs had bones with reduced ash, Ca and P content (P < 0.05). The PhyG at 250 FTU/kg improved bone ash vs. NC. Increasing PhyG dose linearly or quadratically improved bone ash, ADG and FCR (P < 0.05). At ≥ 500 FTU/kg, both PhyG and PhyB maintained ADG and FCR equivalent to PC. Linear regression analysis was done to compare the measured response parameters to increasing digestible P from MCP. Based on this analysis it was shown that PhyG and PhyB at 1,000 FTU/kg could replace 1.83 and 1.66 g/kg digestible P from MCP in the diet, respectively, on average across metacarpi bone ash, ADG or FCR. These findings suggest that the biosynthetic phytase is highly effective in the tested dietary setting.