Suscripción a Biblioteca: Guest
Portal Digitalde Biblioteca Digital eLibros Revistas Referencias y Libros de Ponencias Colecciones
Critical Reviews™ in Eukaryotic Gene Expression
Factor de Impacto: 2.156 Factor de Impacto de 5 años: 2.255 SJR: 0.649 SNIP: 0.599 CiteScore™: 3

ISSN Imprimir: 1045-4403
ISSN En Línea: 2162-6502

Volumen 30, 2020 Volumen 29, 2019 Volumen 28, 2018 Volumen 27, 2017 Volumen 26, 2016 Volumen 25, 2015 Volumen 24, 2014 Volumen 23, 2013 Volumen 22, 2012 Volumen 21, 2011 Volumen 20, 2010 Volumen 19, 2009 Volumen 18, 2008 Volumen 17, 2007 Volumen 16, 2006 Volumen 15, 2005 Volumen 14, 2004 Volumen 13, 2003 Volumen 12, 2002 Volumen 11, 2001 Volumen 10, 2000 Volumen 9, 1999 Volumen 8, 1998 Volumen 7, 1997 Volumen 6, 1996 Volumen 5, 1995 Volumen 4, 1994

Critical Reviews™ in Eukaryotic Gene Expression

DOI: 10.1615/CritRevEukaryotGeneExpr.2020034410
pages 411-425

Molecular Cloning and Characterization of an Acidic Polygalacturonase from Grapes and Its Potential in Industry

Zahra Nazir
Institute of Biochemistry and Biotechnology, University of the Punjab, Lahore, Pakistan
Riffat Chohan
Institute of Biochemistry and Biotechnology, University of the Punjab, Lahore, Pakistan
Malik Siddique Mahmood
Institute of Biochemistry and Biotechnology, University of the Punjab, Lahore, Pakistan
Hafiz Muzzammel Rehman
Institute of Biochemistry and Biotechnology, University of the Punjab, Lahore, Pakistan
Roquyya Gul
Institute of Biological Sciences, Gulab Devi Educational Complex, Lahore, Pakistan
Mahjabeen Saleem
Institute of Biochemistry and Biotechnology, University of the Punjab, Lahore, Pakistan


Pectinase enzymes from different plants have many possible biotechnological applications in various industries. Considering industrial significance of pectinolytic enzymes, the polygalacturonase (PG) gene from grapes was cloned into Escherichia coli DH5α using pTZ57R/T vector. Homologous sequences established a close link to Vitis vinifera. Conserve domain analysis predicted PLN02218 domain belongs to the cl31843 superfamily, showing its function as polygalacturonase. After confirmation by PCR and restriction analysis, the PG gene was expressed in E. coli BL21 and induced by IPTG. Expression level was assessed by 12% SDS-PAGE, which showed a 47 kDa protein. High expression level in the soluble fraction indicated that the protein is intracellular or transmembrane. Maximum expression was obtained with 1 mM IPTG and 6 hour induction time, and autoinduction with lactose increased production of the recombinant enzyme. Zymogram analysis revealed that the induced protein was an active enzyme. Expressed PG showed pectinolytic effect on the physiochemical properties of lemon juice. Natural biopolymers are greatly needed because synthetic fibers can negatively affect health. Pectin hydrolysis of banana pseudostem by the PG enzyme produced better quality fiber. Morphological studies by scanning electron microscopy revealed pectin degradation within the fiber cell architecture, showing the effectiveness of PG treatment on banana pseudostem.


  1. Sandri IG, Fontana RC, Barfknecht DM, da Silveir MM. Clarification of fruit juices by fungal pectinases. LWT-Food Sci Technol. 2011;44(10):2217-22.

  2. Tapre AR, Jain RK. Pectinases: Enzymes for fruit processing industry. Int Food Res J. 2014;21(2):447-53.

  3. Kashyap DR, Vohra PK, Chopra S, Tewari R. Applications of pectinases in the commercial sector: A review. Bioresour Technol. 2001;77:215-27. doi: 10.1016/ S0960-8524(00)00118-8.

  4. Endler A, Persson S. Cellulose synthases and synthesis in Arabidopsis. Mol Plant. 2011;4(2):199-211.

  5. Willats WG, Knox JP, Mikkelsen JD. Pectin: New insights into an old polymer are starting to gel. Trends Food Sci Technol. 2006;17(3):97-104.

  6. Jayani RS, Saxena S, Gupta R. Microbial pectinolytic enzymes: A review. Process Biochem. 2005;40:2931-44. doi: 10.1016/j.procbio.2005.03.026.

  7. Pedrolli DB, Monteiro AC, Gomes E, Carmona EC. Pectin and pectinases: Production, characterization and industrial application of microbial pectinolytic enzymes. Open Bio-technol J. 2009;3:9-18.

  8. Jach G, SoezerN, Schullehner K, Lalla B, Welters P, Muelle A. Phytomining of plant enzymes for biotechnological use of fats and oils. Eur J Lipid Sci Technol. 2010;112:75-86. doi: 10.1002/ejlt.200900100.

  9. M'Hamdi N, Darej C, Jebali J, Bouraoui R, Metahni S, Frouja I, Singh DG, Jarboui I, Brar SK. Different enzymes and their production. In: Brar SK, Verma M, editors. Enzymes in value-addition of wastes. New York: Nova; 2014. p. 109-32.

  10. Khan AW, Shafiq T, Ahmad M. Physical and biochemical changes in commonly grown grapes (Vitis vinifera) in Pakistan at different maturity levels. Pak J Sci. 2008;60:3-4.

  11. Osorio JCM, Baracaldo RB, Florez JJO. The influence of alkali treatment on banana fibre's mechanical properties. Ing Inv. 2012;32(1):83-7.

  12. Reddy KO, Maheswari CU, Shukla M, Song JI, Rajulu AV. Tensile and structural characterization of alkali treated Borassus fruit fine fibers. Compos B Eng. 2013;44(1):433-8.

  13. Wong DW. The ABCs of gene cloning. New York: Springer; 2006.

  14. Sambrook J, Russell DW. Molecular cloning: A laboratory manual. New York: Cold Spring Harbor Laboratory Press; 2001.

  15. Hall TA. BioEdit: A user-friendly biological sequence alignment editor and analysis program for Windows 95/98/NT. Nucleic Acids Symp Ser. 1999;41:95-8.

  16. Gasteiger E, Gattiker A, Hoogland C, Ivanyi I, Appel RD, Bairoch A. ExPASy: The proteomics server for indepth protein knowledge and analysis. Nucleic Acid Res. 2003;31(13):3784-8.

  17. Kallberg M, Wang H, Wang S, Peng J, Wang Z, Lu H, Xu J. Template-based protein structure modeling using the RaptorX web server. Nat Protoc. 2012;7(8):1511-22.

  18. Ko CH, Tsai CH, Tu J, Tang SH, Liu CC. Expression and thermostability of Paenibacillus campinasensis BL11 pectate lyase and its applications in bast fibre processing. Ann Appl Bio. 2011;158(2):218-25.

  19. Dassault Systemes BIOVIA. Discovery studio modeling environment. Velizy-Villacoublay, France; 2017.

  20. Laemmli UK. Cleavage of structural proteins during the assembly of the head of bacteriophage T4. Nature. 1970;227(5359):680-5.

  21. Miller GL. Use of dinitrosalicylic acid reagent for determi-nation of reducing sugar. Anal Chem. 1959;31(3):426-8.

  22. Bradford MM. A rapid and sensitive method for the quantitation of microgram quantities of protein utilizing the principle of protein-dye binding. Anal Biochem. 1976;72(1-2):248-54.

  23. Davis BJ. Disc electrophoresis - II method and application to human serum proteins. Ann N Y Acad Sci. 1964;121(2):404-27.

  24. Saxena S, Shukla S, Thakur A, Gupta R. Immobilization of polygalacturonase from Aspergillus niger onto activated polyethylene and its application in apple juice clarification. Acta Microbiol Immunol Hung. 2008;55(1):33-51.

  25. Blommel PG, Becker KJ, Duvnjak P, Fox BG. Enhanced bacterial protein expression during auto-induction obtained by alteration of lac repressor dosage and medium composition. Biotechnol Prog. 2007;23(3):585-98.

  26. Tu T, Meng K, Bai Y, Shi P, Luo H, Wang Y, Yang P, Zhang Y, Zhang W, Yao B. High-yield production of a low-tem-perature-active polygalacturonase for papaya juice clarification. Food Chem. 2013;141(3):2974-81.

  27. Maktouf S, Neifar M, Drira SJ, Baklouti S, Fendri M, Chaabouni SE. Lemon juice clarification using fungal pectinolytic enzymes coupled to membrane ultrafiltration. Food Bioprod Process. 2014;92(1):14-9.

  28. Hyson DA. A review and critical analysis of the scientific literature related to 100% fruit juice and human health. Adv Nutr. 2015;6(1):37-51.

  29. Prasanna V, Prabha TN, Tharanathan RN. Fruit ripening phenomena-an overview. Crit Rev Food Sci Nutr. 2007;47(1):1-19.

  30. Bag A, Bhattacharyya SK, Pal NK, Chattopadhyay RR. In vitro antimicrobial potential of Terminalia chebula fruit extracts against multidrug-resistant uropathogens. Asian Pac J Trop Biomed. 2012;2(3):S1883-7.

  31. Bansode DS, Chavan MD. Evaluation of antimicrobial activity and phytochemical analysis of papaya and pineapple fruit juices against selected enteric pathogens. Int J Pharma Bio Sci. 2013;4(2):B1176-84.

  32. Negi PS. Plant extracts for the control of bacterial growth: Efficacy, stability and safety issues for food application. Int J Food Microbiol. 2012;156(1):7-17.

  33. Shroff A, Karolia A, Shah J. Bio-softening of banana fiber for nonwoven application. Int J Sci Res. 2015;4(4):524-7.

  34. Jacob N, Niladevi KN, Anisha GS, Prema P. Hydrolysis of pectin: an enzymatic approach and its application in banana fiber processing. Microbiol Res. 2008;163(5):538-44.

  35. Sharma DC, Satyanarayana T. A marked enhancement in the production of a highly alkaline and thermostable pectinase by Bacillus pumilus dcsr1 in submerged fermentation by using statistical methods. Bioresour Technol. 2006;97(5):727-33.

  36. Diano N, Grimaldi T, Bianco M, Rossi S, Gabrovska K, Yordanova G, Godjevargova T, Grano V, Nicolucci C, Mita L, Bencivenga U, Canciglia P, Mita DG. Apple juice clarification by immobilized pectolytic enzymes in packed or fluidized bed reactors. J Agric Food Chem. 2008;56(23):11471-7.

  37. Baley C, Morvan C, Grohens Y. Influence of the absorbed water on the tensile strength of flax fibers. Macromol Symp. 2005;222(1):195-202.

  38. Bourmaud A, Morvan C, Baley C. Importance of fiber preparation to optimize the surface and mechanical properties of unitary flax fiber. Ind Crops Prod. 2010;32(3):662-7.

Articles with similar content:

1st World Congress of Young Scientists on Hydrogen Energy Systems, Vol.0, 2005, issue
Chong Zhang, Xin-Hui Xing, Yun-Li Ren
Optimization of Fermentation Process and Its Impact on Gene Transcription of Intracellular Polysaccharide Synthesis in the Wood Ear Medicinal Mushroom Auricularia auricula-judae (Agaricomycetes)
International Journal of Medicinal Mushrooms, Vol.22, 2020, issue 6
Hong Gao, Chao-Min Yin, De-Fang Shi, Xiu-Zhi Fan, Fen Yao
Farming Technology, Biochemistry Characterization, and Protective Effects of Culinary-Medicinal Mushrooms Agaricus brasiliensis S.Wasser et al. and Lentinus edodes (Berk.) Singer: Five Years of Research in Brazil
International Journal of Medicinal Mushrooms, Vol.7, 2005, issue 1&2
Daisy Maria Favero Salvadori, Lucia Regina Ribeiro, Robson M. Di Piero, Luis Fernando Barbisan, Sergio Florentino Pascholati, Augusto Ferreira da Eira, Patricia Lepage Alves de Lima, Ramon Kaneno, Edson Rodrigues Filho
Cell Cycle and Cancer: Critical Events at the G1 Restriction Point
Critical Reviews™ in Oncogenesis, Vol.7, 1996, issue 1-2
Michele Pagano, Massimo Loda, Giannino DelSal
Development of an Expression Plasmid and its Use in Genetic Manipulation of Lingzhi or Reishi Medicinal Mushroom, Ganoderma lucidum (Higher Basidiomycetes)
International Journal of Medicinal Mushrooms, Vol.16, 2014, issue 2
Meng-Fei Ren, Jun-Wei Xu, Yi-Long He, Xuya Yu, Sen-Lin Ji