A novel amylase-based biocatalyst developed by researchers at the Central Leather Research Institute (CSIR-CLRI), Chennai, helps in processing leather in an environment-friendly way and also drastically cutting the time taken to process the skin at the pre-tanning stage. Pre-tanning process generates 60-70% of total pollution during processing.
Reduced effluents
The quantum of effluent discharge is considerably cut as there is threefold reduction in water usage when the biocatalyst is used. In particular, the amount of chromium that gets absorbed is more leading to less chromium discharge into the environment. Chromium is used for increasing the stability of the collagen through cross-linking. Since no chemicals are used, the chemical oxygen demand drops by about 35% while the total solid effluent load reduces by over 50%.
The reason why less chromium and water are required at the pre-tanning stage when the biocatalyst is used is primarily because of the 120-fold higher binding of the biocatalyst to the glycan sugar (glycosaminoglycan) present predominantly in the skin. Once the catalysts binds to the sugar, it selectively breaks down (hydrolysis) the sugar thus opening up the skin fibre. The results were published in the journal Green Chemistry.
Quick hydrolysis
“The binding and hydrolysis happens rapidly. It takes just 10 minutes to open the fibres when the biocatalyst that we engineered is used. Traditionally, enzymes take three-to-four hours to open the fibres. If lime and sulphate are used it takes 12 hours to complete the process,” says Niraikulam Ayyadurai from the Department of Biochemistry and Biotechnology at CLRI and corresponding author of the paper.
Not only is the process of opening the fibres quicker, the biocatalyst also penetrates deep into the skin unlike the traditionally used enzymes. Deep penetration of the biocatalyst has two advantages — it is sufficient to use less amount of chromium to increase the stability of the collagen and the quality of the finished leather also becomes superior. About 21% of the chromium used gets absorbed by the skin, which is far more than when other enzymes or chemical-based methods are used, leading to reduced chromium in the effluent discharge.
The biocatalyst is stable even at a high temperature of 90 degree C and pH 10 and so up to 95% of the enzyme can be recovered after a single process and reused.
Genetic engineering
The normal amylase enzyme has limited efficiency to bind to the substrate leading to reduced ability to open the skin fibre. So the team led by Dr. Ayyadurai resorted to genetic code. “The genetic code engineering allows us to introduce new chemistry in the amylase enzyme thus improving the enzymatic properties,” he says.
“The tyrosine amino acid was computationally modified with extra groups such as amino, hydroxyl, fluorine and chlorine. We found the extra hydroxyl group provided more activity towards the skin glycan,” says Suryalakshmi Pandurangan from CLRI and the first author of the paper. “By modifying the tyrosine amino acid we changed the property of amylase enzyme.”
The amylase gene was isolated from Bacillus licheniformis and made to express in E. coli. Large-scale production and even manipulation of the enzyme is possible when the enzyme is expressed by E. coli. It also becomes cheaper to produce the enzyme through the fermentation route.
