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N∆89 and C∆274 Truncated Enzymes of Chondroitinase ABC I Regain More Imperturbable Microenvironments Around Structural Components in Comparison to their Wild Type

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Abstract

Immune response stimulation and inactivation of chondroitinase ABC I in physiological condition have been limited its use in various clinical conditions as a bacterial enzyme drug. In the present study, we have investigated some structural and functional features of N∆89, C∆274 and N∆89C∆274; three designed truncated cABC I, in order to clarify the unclear role of two terminal parts of cABC I i.e., the 1–89 and 747–1021 amino acids sequences of the full length enzyme through truncation. As a result, the numbers of potential epitopes, the susceptibility to trypsin digestion, ANS fluorescence spectra, and fluorescence quenching using KI and acrylamide were diminished for N∆89 and C∆274 in comparison to the wild type. Secondary and tertiary structure investigation for N∆89 and C∆274 revealed that the intrinsic fluorescence was increased and Far-UV CD spectra were changed accordingly. Relative to the wild type enzyme, 0.164, 0.195 remaining activity and lack of activity was shown with the zymographic assay for N∆89, C∆274 and N∆89C∆274 variants, respectively. The diminished enzyme activity and structural changes suggested a reorientation of microenvironments interactions including cation–π interactions around structural elements toward lowering regional mobility. Constructing applicable truncated cABC I with improved features could be regarded as a strategy to regain new possible functional advantages over the full length enzyme.

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Abbreviations

cABC I:

Chondroitinase ABC I

cAC:

Chondroitinase AC

PIC:

Protein interaction calculator

LB:

Luria–Bertani

IPTG:

Isopropyl-ß-d-thiogalactopyranoside

PMSF:

Phenylmethanesulfonyl fluoride

DAB:

3,3′-Diaminobenzidine

SDS-PAGE:

Sodium dodecyl sulfate-polyacrylamide gel electrophoresis

ANS:

8-Anilinonaphthalene-1-sulfonic acid

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Acknowledgements

Financial support for this work was provided by Research Council of Tehran University of Medical Sciences under Contract Number of 29977.

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Authors and Affiliations

  1. Department of Clinical Biochemistry, School of Medicine, Tehran University of Medical Sciences, P.O. Box: 14155–6447, Tehran, Iran

    Hossein Omidi-Ardali, Mahdi Aminian, Abolfazl Golestani & Monireh Maleki

  2. Research Center for Biochemistry and Nutrition in Metabolic Diseases, Kashan University of Medical Sciences, Kashan, Iran

    Mohammad Esmaeil Shahaboddin

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  1. Hossein Omidi-Ardali
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  2. Mahdi Aminian
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  3. Abolfazl Golestani
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Correspondence to Mahdi Aminian or Abolfazl Golestani.

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Omidi-Ardali, H., Aminian, M., Golestani, A. et al. N∆89 and C∆274 Truncated Enzymes of Chondroitinase ABC I Regain More Imperturbable Microenvironments Around Structural Components in Comparison to their Wild Type. Protein J 38, 151–159 (2019). https://doi.org/10.1007/s10930-019-09821-y

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