Azo dying of α-keratin material improves microbial keratinase screening and standardization
Research output: Contribution to journal › Journal article › Research › peer-review
Standard
Azo dying of α-keratin material improves microbial keratinase screening and standardization. / Gonzalo, Milena; Espersen, Roall; Al-Soud, Waleed A.; Cristiano Falco, Francesco; Hägglund, Per; Sørensen, Søren J.; Svensson, Birte; Jacquiod, Samuel.
In: Microbial Biotechnology, Vol. 13, No. 4, 2020, p. 984-996.Research output: Contribution to journal › Journal article › Research › peer-review
Harvard
APA
Vancouver
Author
Bibtex
}
RIS
TY - JOUR
T1 - Azo dying of α-keratin material improves microbial keratinase screening and standardization
AU - Gonzalo, Milena
AU - Espersen, Roall
AU - Al-Soud, Waleed A.
AU - Cristiano Falco, Francesco
AU - Hägglund, Per
AU - Sørensen, Søren J.
AU - Svensson, Birte
AU - Jacquiod, Samuel
PY - 2020
Y1 - 2020
N2 - Microbial conversion through enzymatic reactions has received a lot of attention as a cost-effective and environmentally friendly way to recover amino acids and short peptides from keratin materials. However, accurate assessment of microbial keratinase activity is not straightforward, and current available methods lack sensitivity and standardization. Here, we suggest an optimized Azokeratin assay, with substrate generated directly from azo-dyed raw keratin material. We introduced supernatant filtration in the protocol for optimal stopping of keratinase reactions instead of the widely used trichloroacetic acid (TCA), as it generated biases and impacted the sensitivity. We furthermore suggest a method for standardization of keratinase activity signals using proteinase K, a well-known keratinase, as a reference enabling reproducibility between studies. Lastly, we evaluated our developed method with several bacterial isolates through benchmarking against a commercial assay (Keratin Azure). Under different setups, the Azokeratin method was more sensitive than commonly used Keratin Azure-based assays (3-fold). We argue that this method could be applied with any type of keratin substrate, enabling more robust and sensitive results which can be used for further comparison with other studies, thus representing an important progress within the field of microbial keratin degradation.
AB - Microbial conversion through enzymatic reactions has received a lot of attention as a cost-effective and environmentally friendly way to recover amino acids and short peptides from keratin materials. However, accurate assessment of microbial keratinase activity is not straightforward, and current available methods lack sensitivity and standardization. Here, we suggest an optimized Azokeratin assay, with substrate generated directly from azo-dyed raw keratin material. We introduced supernatant filtration in the protocol for optimal stopping of keratinase reactions instead of the widely used trichloroacetic acid (TCA), as it generated biases and impacted the sensitivity. We furthermore suggest a method for standardization of keratinase activity signals using proteinase K, a well-known keratinase, as a reference enabling reproducibility between studies. Lastly, we evaluated our developed method with several bacterial isolates through benchmarking against a commercial assay (Keratin Azure). Under different setups, the Azokeratin method was more sensitive than commonly used Keratin Azure-based assays (3-fold). We argue that this method could be applied with any type of keratin substrate, enabling more robust and sensitive results which can be used for further comparison with other studies, thus representing an important progress within the field of microbial keratin degradation.
U2 - 10.1111/1751-7915.13541
DO - 10.1111/1751-7915.13541
M3 - Journal article
C2 - 32110845
VL - 13
SP - 984
EP - 996
JO - Microbial Biotechnology
JF - Microbial Biotechnology
SN - 1751-7907
IS - 4
ER -
ID: 237651979