Abstract
In this study, we report a chromogenic reaction between magnesium ascorbyl phosphate (MAP) and ferric chloride to generate a Brown-Red clathrate, while the Treated MAP by phosphatases forms Colorless (BRTC) product with ferric chloride. The BRTC was indicative of phosphatase activity-mediated excision of phosphorous group from MAP and utilized to screen phosphatases from bacterial cell lysates. From ten tested strains, BRTC was observed in the cell lysate of Salmonella enterica subsp. enterica serovar Cerro 87. BRTC was again employed to track phosphatase activity of the resuspensions of the ammonium sulfate graded precipitations of the cell lysate. Two phosphatases, PhoN and YcdX, were identified by LC-MS/MS analysis in the protein fraction giving most obvious BRTC phenotype and validated by examination of in vitro activity of the purified proteins.
Key points
• BRTC is labelling-free, naked-eye visible, and independent of any facilities.
• BRTC can directly screen phosphatases from microbial cell lysates.
• Using BRTC system, two phosphatases were identified in Salmonella enterica subsp. enterica serovar Cerro 87.
Graphical abstract
This is a preview of subscription content, log in via an institution to check access.
Similar content being viewed by others
Data availability
The data supporting the findings of this study are available within the article and its supplementary information files.
References
Abedini F, Hosseinkhani H, Ismail M, Domb AJ, Omar AR, Chong PP, Hong PD, Yu DS, Farber IY (2012) Cationized dextran nanoparticle-encapsulated CXCR4-siRNA enhanced correlation between CXCR4 expression and serum alkaline phosphatase in a mouse model of colorectal cancer. Int J Nanomedicine 7:4159–4168. https://doi.org/10.2147/ijn.S29823
Ahmadzadeh E, Talebnia F, Tabatabaei M, Ahmadzadeh H, Mostaghaci B (2016) Osteoconductive composite graft based on bacterial synthesized hydroxyapatite nanoparticles doped with different ions: from synthesis to in vivo studies. Nanomedicine 12(5):1387–1395. https://doi.org/10.1016/j.nano.2016.01.020
Amr S, Brown MB, Martin GP, Forbes B (2001) Activation of clindamycin phosphate by human skin. J Appl Microbiol 90(4):550–554. https://doi.org/10.1046/j.1365-2672.2001.01282.x
Avonce N, Mendoza-Vargas A, Morett E, Iturriaga G (2006) Insights on the evolution of trehalose biosynthesis. BMC Evol Biol 6:109. https://doi.org/10.1186/1471-2148-6-109
Bautista FM, Bravo MC, Campelo JM, Garcia A, Luna D, Marinas JM, Romero AA (1999) Covalent immobilization of acid phosphatase on amorphous AlPO4 support. J Mol Catal B Enzym 6(5):473–481. https://doi.org/10.1016/S1381-1177(99)00005-3
Behera BC, Yadav H, Singh SK, Mishra RR, Sethi BK, Dutta SK, Thatoi HN (2017a) Phosphate solubilization and acid phosphatase activity of Serratia sp. isolated from mangrove soil of Mahanadi river delta, Odisha, India. J. Genet. Eng. Biotechnol 15(1):169–178. https://doi.org/10.1016/j.jgeb.2017.01.003
Behera BC, Yadav H, Singh SK, Sethi BK, Mishra RR, Kumari S, Thatoi H (2017b) Alkaline phosphatase activity of a phosphate solubilizing Alcaligenes faecalis, isolated from Mangrove soil. Biotechnol Res Innovation 1(1):101–111. https://doi.org/10.1016/j.biori.2017.01.003
Browning TJ, Achterberg EP, Yong JC, Rapp I, Utermann C, Engel A, Moore CM (2017) Iron limitation of microbial phosphorus acquisition in the tropical North Atlantic. Nat Commun 8:15465. https://doi.org/10.1038/ncomms15465
Calderone V, Forleo C, Benvenuti M, Cristina Thaller M, Rossolini GM, Mangani S (2004) The first structure of a bacterial class B Acid phosphatase reveals further structural heterogeneity among phosphatases of the haloacid dehalogenase fold. J Mol Biol 335(3):761–773. https://doi.org/10.1016/j.jmb.2003.10.050
Chang M-Y, Juang R-S (2007) Stability and reactivity of acid phosphatase immobilized on composite beads of chitosan and ZrO2 powders. Int J Biol Macromol 40(3):224–231. https://doi.org/10.1016/j.ijbiomac.2006.07.009
Chaudhuri G, Chatterjee S, Venu-Babu P, Ramasamy K, Thilagaraj WR (2013) Kinetic behaviour of calf intestinal alkaline phosphatase with pNPP. Indian J Biochem Biophys 50(1):64–71
Coleman JE (1992) Structure and mechanism of alkaline phosphatase. Annu Rev Biophys Biomol Struct 21:441–483. https://doi.org/10.1146/annurev.bb.21.060192.002301
Danzer T, Schwedt G (1996) Chemometric methods for the development of a biosensor system and the evaluation of inhibition studies with solutions and mixtures of pesticides and heavy metals Part 1. Development of an enzyme electrodes system for pesticide and heavy metal screening using selected chemometric methods. Anal Chim Acta 318(3):275–286. https://doi.org/10.1016/0003-2670(95)00460-2
Fei Y, Huang S, Zhang H, Tong Y, Wen D, Xia X, Wang H, Luo Y, Barceló D (2020) Response of soil enzyme activities and bacterial communities to the accumulation of microplastics in an acid cropped soil. Sci Total Environ 707:135634. https://doi.org/10.1016/j.scitotenv.2019.135634
Gangappa R, Yong P, Singh S, Mikheenko IJ, Murray A, Macaskie L (2016) Hydroxyapatite biosynthesis by a Serratia sp. and application of nanoscale bio-HA in the recovery of strontium and europium. Geomicrobiology 33:267–273. https://doi.org/10.1080/01490451.2015.1067657
Green MR, Sambrook J (2020) Dephosphorylation of DNA fragments with alkaline phosphatase. Cold Spring Harb Protoc 2020(8):100669. https://doi.org/10.1101/pdb.prot100669
Han C, Wang Z, Wu Q, Yang W, Yang H, Xue X (2016) Evaluation of the role of inherent Ca2+ in phosphorus removal from wastewater system. Water Sci Technol 73(7):1644–1651. https://doi.org/10.2166/wst.2015.642
He W, Huang T, Tang Y, Liu Y, Wu X, Chen S, Chan W, Wang Y, Liu X, Chen S, Wang L (2015) Regulation of DNA phosphorothioate modification in Salmonella enterica by DndB. Sci Rep 5:12368. https://doi.org/10.1038/srep12368
Hulett FM, Kim EE, Bookstein C, Kapp NV, Edwards CW, Wyckoff HW (1991) Bacillus subtilis alkaline phosphatases III and IV. Cloning, sequencing, and comparisons of deduced amino acid sequence with Escherichia coli alkaline phosphatase three-dimensional structure. J Biol Chem 266(2):1077–1084
Kaliannan K, Hamarneh SR, Economopoulos KP, Nasrin Alam S, Moaven O, Patel P, Malo NS, Ray M, Abtahi SM, Muhammad N, Raychowdhury A, Teshager A, Mohamed MM, Moss AK, Ahmed R, Hakimian S, Narisawa S, Millán JL, Hohmann E, Warren HS, Bhan AK, Malo MS, Hodin RA (2013) Intestinal alkaline phosphatase prevents metabolic syndrome in mice. Proc Natl Acad Sci U S A 110(17):7003–7008. https://doi.org/10.1073/pnas.1220180110
Kulkarni S, Chin E, Tran V, Ho MKM, Phan C, Sommerhalter M (2010) Sol–gel encapsulation of acid phosphatase in the presence of the ionic liquid [BMIM] [BF4]. Monatshefte für Chemie - Chemical Monthly 141(1):119–123. https://doi.org/10.1007/s00706-009-0230-7
Kurita K, Yoshino H, Nishimura S-I, Ishii S, Mori T, Nishiyama Y (1997) Mercapto-chitins: a new type of supports for effective immobilization of acid phosphatase. Carbohydr Polym 32(3):171–175. https://doi.org/10.1016/S0144-8617(97)00010-6
Lee SW, Oh MK (2016) Improved production of N-acetylglucosamine in Saccharomyces cerevisiae by reducing glycolytic flux. Biotechnol Bioeng 113(11):2524–2528. https://doi.org/10.1002/bit.26014
Makde RD, Mahajan SK, Kumar V (2007) Structure and mutational analysis of the PhoN protein of Salmonella typhimurium provide insight into mechanistic details. Biochemistry 46(8):2079–2090. https://doi.org/10.1021/bi062180g
McComb RB, Bowers GN Jr (1985) Alkaline phosphatase and the International Clinical Enzyme Scale. Am J Clin Pathol 84(1):67–73. https://doi.org/10.1093/ajcp/84.1.67
Meng D, Liang A, Wei X, You C (2019) Enzymatic characterization of a thermostable phosphatase from Thermomicrobium roseum and its application for biosynthesis of fructose from maltodextrin. Appl Microbiol Biotechnol 103(15):6129–6139. https://doi.org/10.1007/s00253-019-09917-6
Muniyan S, Chaturvedi NK, Dwyer JG, Lagrange CA, Chaney WG, Lin MF (2013) Human prostatic acid phosphatase: structure, function and regulation. Int J Mol Sci 14(5):10438–10464. https://doi.org/10.3390/ijms140510438
Ou L, Huang B, Hong H, Qi Y, Lu S (2010) Comparative alkaline phosphatase characteristics of the algal bloom dinoflagellates Prorocentrum donghaiense and Alexandrium catenella, and the diatom Skeletonema costatum. J Phycol 46(2):260–265. https://doi.org/10.1111/j.1529-8817.2009.00800.x
Posen S (1967) Alkaline phosphatase. Ann Intern Med 67(1):183–203. https://doi.org/10.7326/0003-4819-67-1-183
Rankin SA, Christiansen A, Lee W, Banavara DS, Lopez-Hernandez A (2010) Invited review: the application of alkaline phosphatase assays for the validation of milk product pasteurization. J Dairy Sci 93(12):5538–5551. https://doi.org/10.3168/jds.2010-3400
Rossolini GM, Schippa S, Riccio ML, Berlutti F, Macaskie LE, Thaller MC (1998) Bacterial nonspecific acid phosphohydrolases: physiology, evolution and use as tools in microbial biotechnology. Cell Mol Life Sci 54(8):833–850. https://doi.org/10.1007/s000180050212
Segundo-Val IS, Sanz-Lozano CS (2016) Introduction to the gene expression analysis. Methods Mol Biol 1434:29–43. https://doi.org/10.1007/978-1-4939-3652-6_3
Thistleton J, Berry TA, Pearce P, Parsons SA (2002) Mechanisms of chemical phosphorus removal II: iron (III) salts. Process Saf Environ Prot 80(5):265–269. https://doi.org/10.1205/095758202762277623
Wang J, Stieglitz KA, Kantrowitz ER (2005) Metal specificity is correlated with two crucial active site residues in Escherichia coli alkaline phosphatase. Biochemistry 44(23):8378–8386. https://doi.org/10.1021/bi050155p
Wang E, Koutsioulis D, Leiros HK, Andersen OA, Bouriotis V, Hough E, Heikinheimo P (2007) Crystal structure of alkaline phosphatase from the Antarctic bacterium TAB5. J Mol Biol 366(4):1318–1331. https://doi.org/10.1016/j.jmb.2006.11.079
Wojciechowski CL, Cardia JP, Kantrowitz ER (2002) Alkaline phosphatase from the hyperthermophilic bacterium T. maritima requires cobalt for activity. Protein Sci 11(4):903–911. https://doi.org/10.1110/ps.4260102
Xu X, Wozniczka M, Van Hecke K, Buyst D, Mara D, Vervaet C, Herman K, Wynendaele E, Deconinck E, De Spiegeleer B (2020) Structural study of L-ascorbic acid 2-phosphate magnesium, a raw material in cell and tissue therapy. J Biol Inorg Chem 25(6):875–885. https://doi.org/10.1007/s00775-020-01801-3
Yang H, He K, Lu D, Wang J, Xu D, Jin Z, Yang M, Chen J (2020) Removal of phosphate by aluminum-modified clay in a heavily polluted lake, Southwest China: effectiveness and ecological risks. Sci Total Environ 705:135850. https://doi.org/10.1016/j.scitotenv.2019.135850
You C, Shi T, Li Y, Han P, Zhou X, Zhang YP (2017) An in vitro synthetic biology platform for the industrial biomanufacturing of myo-inositol from starch. Biotechnol Bioeng 114(8):1855–1864. https://doi.org/10.1002/bit.26314
Zhang WH, Sun RB, Xu L, Liang JN, Zhou J (2019) Assessment of bacterial communities in Cu-contaminated soil immobilized by a one-time application of micro-/nano-hydroxyapatite and phytoremediation for 3 years. Chemosphere 223:240–249. https://doi.org/10.1016/j.chemosphere.2019.02.049
Zhu J, Huang Q, Pigna M, Violante A (2010) Immobilization of acid phosphatase on uncalcined and calcined Mg/Al-CO3 layered double hydroxides. Colloids Surf. B Biointerfaces 77(2):166–173. https://doi.org/10.1016/j.colsurfb.2010.01.020
Funding
This work was supported by grant from the National Key Research and Development Program of China (grant number 2020YFA0907200) and the National Natural Science Foundation of China (grant numbers 31871250, 31670034).
Author information
Authors and Affiliations
Contributions
YW, WH, and XH conceived and designed research. ZD and XH supervised the project. YW conducted experiments. YW and WH analyzed data. YW and XH wrote the manuscript. All authors read and approved the manuscript.
Corresponding author
Ethics declarations
Ethics approval
This article does not contain any studies with human participants or animals performed by any of the authors.
Conflict of interest
The authors declare no competing interests.
Additional information
Publisher’s note
Springer Nature remains neutral with regard to jurisdictional claims in published maps and institutional affiliations.
Rights and permissions
About this article
Cite this article
Wang, Y., Hu, W., Deng, Z. et al. Rapid identification of magnesium ascorbyl phosphate utilizing phosphatase through a chromogenic change-coupled activity assay. Appl Microbiol Biotechnol 105, 2901–2909 (2021). https://doi.org/10.1007/s00253-021-11229-7
Received:
Revised:
Accepted:
Published:
Issue Date:
DOI: https://doi.org/10.1007/s00253-021-11229-7