Purification and Characterization of Exo-Inulinase from Paenibacillus sp. d9 Strain.

Characterization; Enzyme purification; Exo-inulinase; Paenibacillus sp.; Phylogenetic tree; Bacterial Proteins; Glycoside Hydrolases; inulinase; Bacterial Proteins/chemistry; Bacterial Proteins/genetics; Bacterial Proteins/isolation & purification; Glycoside Hydrolases/genetics; Glycoside Hydrolases/isolation & purification; Hydrogen-Ion Concentration; Paenibacillus/enzymology; Paenibacillus/genetics; Phylogeny; Paenibacillus; Analytical Chemistry; Bioengineering; Biochemistry; Organic Chemistry

Abstract :

[en] This study intended to purify and characterise exo-inulinase of diesel-degrading Paenibacillus sp. D9. The whole genome sequencing of Paenibacillus sp. D9 revealed to possess the sacC gene that is encoded as exo-inulinase/levanase. This isolate was capable of producing a maximum of 50.9 IU/mL of exo-inulinase activity within 3 days at 30 °C, 200 rpm and pH of 7.0 on minimal salt medium agar supplemented with 1% (w/v) inulin. An exo-inulinase of 58.5 kDa was purified using ammonium sulphate precipitation, HiTrap QFF column and MMC column chromatographies with a specific activity of 4333 IU/mg, 7.1% recovery and a 4.3-fold increase in purity. The purified D9 exo-inulinase had temperature and pH optimum at 40 °C and pH 4.0, respectively, with the Michaelis constant of 5.5 mM and a maximal velocity of 476.2 IU/mg, respectively. Catalytic constant, k cat was calculated to be 42.6 s-1 with a catalytic efficiency (k cat /K m ) of 7.6 s-1 mM-1. The presence of Ca2+ enhanced the activity of D9 exo-inulinase while Hg2+ completely inhibited the activity, other compounds such as Fe3+ and Cu2+ had an inhibitory effect. The results of amino acid alignment and the complete degradation of inulin into fructose by the purified enzyme confirmed that inulinase from Paenibacillus sp. D9 is an exo-form. The phylogenetic tree based on the protein sequences indicates that bacterial exo-inulinases possess a common ancestry.

Disciplines :

Biochemistry, biophysics & molecular biology

Author, co-author :

Jeza, S; Discipline of Microbiology, School of Life sciences, University of KwaZulu-Natal (Westville Campus), Private Bag X54001, Durban, 4001, South Africa

Maseko, Sibusiso Bonginkhost ; Discipline of Microbiology, School of Life sciences, University of KwaZulu-Natal (Westville Campus), Private Bag X54001, Durban, 4001, South Africa

Lin, J ; Discipline of Microbiology, School of Life sciences, University of KwaZulu-Natal (Westville Campus), Private Bag X54001, Durban, 4001, South Africa. linj@ukzn.ac.za

Language :

English

Title :

Purification and Characterization of Exo-Inulinase from Paenibacillus sp. d9 Strain.

Publication date :

February 2018

Journal title :

Protein Journal

ISSN :

1572-3887

Publisher :

Springer Science and Business Media, LLC, Netherlands

Volume :

Issue :

Pages :

70 - 81

Peer reviewed :

Peer Reviewed verified by ORBi

Additional URL :

https://link.springer.com/article/10.1007/s10930-017-9752-8/fulltext.html

Funding text :

The authors would like to thank the National Research Foundation, South Africa, for providing the bursary during the study.

Available on ORBi :

since 05 January 2026

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