Effect of Poly-Histidine Tag Position toward Inhibition Activity of Secretory Leukocyte Protease Inhibitor as Candidate for Material Wound Healing

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Munadziroh Elly; Ulfa Evi Umayah; Labiqah Amaliah; Asmarani One; Tri Puspaningsih Ni Nyoman

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Journal: Avicenna Journal of Medical Biotechnology Year:2020 | Volume:12 | Issue:1 Page(s): 32-36

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Title

Effect of Poly-Histidine Tag Position toward Inhibition Activity of Secretory Leukocyte Protease Inhibitor as Candidate for Material Wound Healing

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Keywords

Escherichia coli

Gene expression

Poly histidine

Secretory leukocyte protease inhibitor

Abstract

Background: The Secretory leukocyte protease inhibitors (SLPI) has many biological functions including anti-bacterial, anti-fungal, anti-viral, anti-inflammatory, and immuno-modulatory. Previous studies have shown that gene-encoding human SLPI have successfully been expressed in Escherichia coli (E. coli) with a C-terminal polyhistidine tag (His-tag). The aim of this research was to investigate the inhibition activity of N-terminal His-tag position (NSLPI) and C-terminal His-tag position (CSLPI). We hypothesized that a His-tag close to an active site SLPI domain may interfere with the inhibition activity of SLPIs. Methods: A NSLPI and CSLPI were constructed with polymerase chain reaction (PCR) amplification. The PCR products were then ligated into pET-30a plasmid and transformed into E. coli TOP10. Recombinant plasmids were verified by using restriction analysis and nucleotide sequence analysis. pET-NSLPI and pET-CSLPI were then subcloned in E. coli BL21(DE3) for its expression. The SLPI protein was expressed using Isopropyl β-D-1-thiogalactopyranoside induction (IPTG). The inhibition effect of both SLPI against Porcine Pancreatic Elastase (PPE) enzyme was tested using the Nsuccinyil-alanyl-L-alanyl-L-prolyl-L-phenylalanyl-4-nitroanalide (NPN) substrate. Results: The SLPI gene was successfully cloned and expressed in E. coli BL21. Fusion proteins of NSLPI and CSLPI were generated with His-tag in the N-terminal and Cterminal position, respectively. The inhibition effect of NSLPI and CSLPI on PPE indicated that both SLPI were active. The inhibition activity of NSLPI was 66. 7%, higher than CSLPI by 44. 4%. Conclusion: The His-tag position on the C-terminal of SLPI reduced the inhibition activity of SLPI.

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APA: Copy

Munadziroh, Elly, Ulfa, Evi Umayah, Labiqah, Amaliah, Asmarani, One, & Tri Puspaningsih, Ni Nyoman. (2020). Effect of Poly-Histidine Tag Position toward Inhibition Activity of Secretory Leukocyte Protease Inhibitor as Candidate for Material Wound Healing. AVICENNA JOURNAL OF MEDICAL BIOTECHNOLOGY (AJMB), 12(1), 32-36. SID. https://sid.ir/paper/314061/en

Vancouver: Copy

Munadziroh Elly, Ulfa Evi Umayah, Labiqah Amaliah, Asmarani One, Tri Puspaningsih Ni Nyoman. Effect of Poly-Histidine Tag Position toward Inhibition Activity of Secretory Leukocyte Protease Inhibitor as Candidate for Material Wound Healing. AVICENNA JOURNAL OF MEDICAL BIOTECHNOLOGY (AJMB)[Internet]. 2020;12(1):32-36. Available from: https://sid.ir/paper/314061/en

IEEE: Copy

Elly Munadziroh, Evi Umayah Ulfa, Amaliah Labiqah, One Asmarani, and Ni Nyoman Tri Puspaningsih, “Effect of Poly-Histidine Tag Position toward Inhibition Activity of Secretory Leukocyte Protease Inhibitor as Candidate for Material Wound Healing,” AVICENNA JOURNAL OF MEDICAL BIOTECHNOLOGY (AJMB), vol. 12, no. 1, pp. 32–36, 2020, [Online]. Available: https://sid.ir/paper/314061/en

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