Urate observed with Microbacterium sp. ZZJ4-1 that 0.3%

oxidase is enzyme conserved in many species ranging from microorganisms to
mammals; however, primates have lost this activity as consequence of
evolutionary gene mutations. This enzyme catalyzes the oxidative reaction that
converts urate to allantoin, a more soluble and easily excreted compound. Due
to this property urate oxidase may be used for both therapeutical and
diagnostic purposes. Many sources of urate oxidase are
available, but commercial production may be hampered by low productivity and
difficulties in protein purification for clinical applications.
Also, high cost is the major disadvantage of treatment by a variety of
recombinant pharmaceutics including urate oxidase. Hence, the development of
more efficient expression systems to produce large amounts of recombinant proteins
very economically is under focus. In this study, a
total of 76 samples with bacterial strains that were able to grow in media
containing uric acid as sole carbon source were screened. Among these samples,
only 17-1 strain with the higher ability to produce uricase based on the
uricase assay data were selected for further studies. This activity was
detected as clear zones accompanying the growth of bacteria in culture Modified
Bennett’s agar medium (0.3% uric acid). In the present study the maximum
uricase production (15.135 U/ml/min) by 17-1 was achieved with 0.3% uric
acid concentration. These results are in agreement with Lotfy et al, who
found 0.3% uric acid enhanced uricase induction (0.45U/ml) by Bacillus
thermocatenulatus. Similar finding was also observed with Microbacterium
sp. ZZJ4-1 that 0.3% uric acid was the best concentration for uricase
production (0.6U/ml), and 24.2U/ml of uricase was induced by P. aeruginosa using
medium containing 0.3% uric acid at 24 hours. According to morphological,
physiological, and biochemical comparison analysis of the characteristics of
isolate 17-1 and other described isolates, 17-1 belongs to the genus Streptomyces.
The identification results were confirmed using 16S rDNA, which is considered a
powerful tool for deducing phylogenetic and evolutionary relationships among
bacteria, archaebacteria, and eukaryotic organisms. The 16S rDNA sequence was
compared to the GenBank database at the NCBI using the BLAST program. According
to the obtained results, 17-1 was identified as Streptomyces bellus,
which is a new uricase producer.  Interestingly,
17-1 was successfully expressed in E. coli BL21 (DE3) with soluble uricase
enzyme activity of …… IU.

purified enzyme also showed a …..-fold increase in its specific activity (……
IU/mg) compared with the crude uricase of 17-1 (15.135 U/ml/min). The molecular
mass of purified recombinant uricase as determined by SDS-PAGE was estimated to
be 38 kDa. Uricases from different sources may have different molecular masses
and amino acid sequences. In another study, the molecular mass of the uricase
was estimated at 34–54 kDa.  In
conclusion, we reported the successful expression and purification of a
recombinant uricase enzyme from streptomyces in E. coli with production of the
enzyme in a soluble and active form.

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