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. 2024 Jan 31;24(1):44.
doi: 10.1186/s12866-024-03192-w.

Purification and characterization of L-arginine deiminase from Penicillium chrysogenum

Hamed M El-Shora  1 , Nessma A El-Zawawy  2 , Mohamed A Abd El-Rheem  2 , Metwally A Metwally  2
Affiliations

Purification and characterization of L-arginine deiminase from Penicillium chrysogenum

Hamed M El-Shora et al. BMC Microbiol. .

Abstract

L-arginine deiminase (ADI, EC 3.5.3.6) hydrolyzes arginine to ammonia and citrulline which is a natural supplement in health care. ADI was purified from Penicillium chrysogenum using 85% ammonium sulfate, DEAE-cellulose and Sephadex G200. ADI was purified 17.2-fold and 4.6% yield with a specific activity of 50 Umg- 1 protein. The molecular weight was 49 kDa. ADI expressed maximum activity at 40oC and an optimum pH of 6.0. ADI thermostability was investigated and the values of both t0.5 and D were determined. Kd increased by temperature and the Z value was 38oC. ATP, ADP and AMP activated ADI up to 0.6 mM. Cysteine and dithiothreitol activated ADI up to 60 µmol whereas the activation by thioglycolate and reduced glutathione (GSH) prolonged to 80 µmol. EDTA, α,α-dipyridyl, and o-phenanthroline inactivated ADI indicating that ADI is a metalloenzyme. N-ethylmaleimide (NEM), N-bromosuccinimide (NBS), butanedione (BD), dansyl chloride (DC), diethylpyrocarbonate (DEPC) and N-acetyl-imidazole (NAI) inhibited ADI activity indicating the necessity of sulfhydryl, tryptophanyl, arginyl, lysyl, histidyl and tyrosyl groups, respectively for ADI catalysis. The obtained results show that ADI from P. chrysogenum could be a potential candidate for industrial and biotechnological applications.

Keywords: Activation; Active groups; Arginine deiminase; Kinetics; P.chrysogenum; Purification.

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Conflict of interest statement

Competing interests. The authors declare no competing interests.

Figures

Fig. 1
Fig. 1
SDS-PAGE of ADI from P. chrysogenum
Fig. 2
Fig. 2
Effect of different substrate concentrations on ADI activity from P. chrysogenum. (A): L-arginine as substrate and (B): Eadie-Hofstee plot
Fig. 3
Fig. 3
Effect of pH and temperature on purified ADI activity from P. chrysogenum. (A): pH and (B): Temperature
Fig. 4
Fig. 4
Thermostability of purified ADI from P. chrysogenum at 50 to 65oC.(A): enzyme activity and (B): relative activity
Fig. 5
Fig. 5
Determination of Z for purified ADI
Fig. 6
Fig. 6
Effect of different concentrations of adenosine compounds on ADI activity from P. chrysogenum
Fig. 7
Fig. 7
Effect of thiol compounds on ADI from P. chrysogenum. (A): Cysteine, (B): DTT, (C): Thioglycolate and (D): GSH
Fig. 8
Fig. 8
Effect of chelating agent compounds on ADI activity from P. chrysogenum. (A): EDTA, (B): α-α-dipyridyl and (C): o-phenanthroline
Fig. 9
Fig. 9
Effect of different modifiers on ADI activity from P. chrysogenum. (A): NEM, (B): NBS, (C): BD, (D): DC, (E): DEPC and (F): NAI
See this image and copyright information in PMC

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