Isolation, Specificity, and Application of β-N-Acetylhexosaminidases from Penicillium crustosum

Round 1

Reviewer 1 Report

The current work deals with the purification and the biochemical characterization of fungal hexosaminidases. As it is it is nicely written, depicted and examplified. In an overall manner it is of good interest to a large scientific audience. It deserves to be reported as it. Only minor points should be corrected, such as N in most sugars.

Author Response

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Reviewer 2 Report

The authors partially purified several β-hexosaminidases from filamentous fungi and investigated their properties. The culture supernatant from filamentous fungi was purified on a cation-exchange chromatography, but the degree of purification is not shown; to calculate parameters such as kcat and to reveal substrate specificity, the enzyme must be of high purity, so it is necessary to show SDS-PAGE that the enzyme is homogeneous. They showed that the enzymes act not only on 4MU-β-GalNAc but also on 2Cl-4NP-beta-Gal, but the possibility that it is caused by a contaminant enzymes cannot be excluded. Finally, the application of hydrolyzing only 4MU-β-GalNAc from a mixture of 4MU-β-GalNAc and 4MU-α-GalNAc to obtain 4MU-α-GalNAc is shown. But it is obvious that β-hexosaminidase does not act on the α anomer, so there is no novelty.

 

Around L170. Show the purity of the enzymes, by using SDS-PAGE.

L227. Since the authors measured the catalytic constant of the overall reaction, k2 should be written as kcat.

Fig. 4. Add the unit on Y-axis. 

Author Response

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Reviewer 3 Report

1. The title is too long, please simplify it.

2.  In abstract, line 13: enzymatic activity; line 16: optimum pH; optimum temperature.

3. line 141. P. crustosum should be italic.

4. Figure 3A: The abscissa value in Figure 3A should use dots instead of commas.

5. Table 2 and 3: The significant numbers in the tables are not uniform.

 

Author Response

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Round 2

Reviewer 2 Report

The enzyme is not completely purified judging from the result of SDS-PAGE, but other improvements have been appropriately corrected.