# Characterization of a λ-Carrageenase Mutant with the Generation of Long-Chain λ-Neocarrageenan Oligosaccharides

## TL;DR

Scientists engineered a mutant enzyme that produces longer-chain λ-carrageenan oligosaccharides, which could be useful in various industries.

## Contribution

A mutant λ-carrageenase was developed that generates longer-chain oligosaccharides with improved yield and altered properties.

## Key findings

- Deleting the pyrroloquinoline quinone-like domain resulted in long-chain oligosaccharides with a polymerization degree of 10–20.
- The mutant enzyme's yield increased five-fold and fewer inclusion bodies were formed during expression.
- The mutant enzyme showed altered optimal temperature (15 °C) and pH (7.0) with a specific activity of 51.59 U/mg.

## Abstract

λ-carrageenan oligosaccharides can be widely applied in the food, pharmaceutical, medicine and cosmetic industries due to their abundant bioactivities, and they are important products for the high-value utilization of λ-carrageenan. However, oligosaccharides with different degrees of polymerization have different properties, and the final products of λ-carrageenase reported so far are mainly λ-neocarrabiose, λ-neocarratetraose and λ-neocarrahexaose without longer-chain oligosaccharides. Further research is consequently required. Herein, a mutant λ-carrageenase was constructed by deleting the pyrroloquinoline quinone-like domain of OUC-CglA derived from Maribacter vaceletii. Interestingly, it was discovered that the majority of final products of the mutant OUC-CglA-DPQQ were long-chain oligosaccharides with a polymerization degree of 10–20, which underwent significant changes compared to that of OUC-CglA. Additionally, without the pyrroloquinoline quinone-like domain, fewer inclusion bodies were produced throughout the expression process, and the yield of the λ-carrageenase increased about five-fold. However, compared to its parental enzyme, significant changes were made to its enzymatic properties. Its optimal temperature and pH were 15 °C and pH 7.0, and its specific activity was 51.59 U/mg. The stability of the enzyme decreased. Thus, it was found that the deleting domain was related to the formation of inclusion bodies, the stability of the enzyme, the activity of the enzyme and the composition of the products.