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Inducible Operons: lac Operon

Inducible Operons: lac Operon

The lac operon in Escherichia coli is a model for understanding inducible gene regulation and metabolic flexibility. It integrates local control by lactose and global regulation through catabolite repression, enabling E. coli to preferentially metabolize glucose when available and switch to lactose utilization when glucose is scarce.Structure and Function of the lac OperonThe lac operon contains three structural genes: lacZ (β-galactosidase), lacY (lactose permease), and lacA...

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Rational Engineering of Cellobiose 2-Epimerase Through Flexible Loop Modulation and Structure-Guided Sequence

Xinyan Mao¹, Hongbin Zhang¹, Chao Hu¹

¹School of Food and Biological Engineering, Hefei University of Technology, Hefei 230009, China.

|February 27, 2026

Summary

Researchers enhanced lactulose production using enzyme engineering of cellobiose 2-epimerase (CsCE). Optimized mutants and conditions achieved high yields and purity for this functional disaccharide.

Keywords:

cellobiose 2-epimerase flexible loop modulation lactulose structure-guided sequence alignment

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Published on: October 24, 2016

Area of Science:

Biotechnology
Enzyme Engineering
Biocatalysis

Background:

Lactulose is a functional disaccharide with significant pharmaceutical and food applications.
Enzymatic isomerization of lactose is an efficient method for lactulose synthesis.

Purpose of the Study:

To enhance lactulose production through protein engineering of cellobiose 2-epimerase (CsCE) and process optimization.
To develop superior CsCE variants with improved activity, stability, and substrate affinity.

Main Methods:

Employed a dual-track protein engineering strategy involving flexible loop modulation and structure-guided sequence alignment.
Created and characterized two superior CsCE mutants: R17Q/L184S and R17Q/S142T.
Optimized reaction conditions including temperature, pH, substrate concentration, and enzyme loading.

Main Results:

The R17Q/L184S mutant showed a 37% increase in activity, enhanced thermostability, and improved substrate affinity.
The R17Q/S142T mutant exhibited the highest specific activity (24.08 U/mg), a 21% increase.
Optimized conditions yielded 75.6% lactulose with >95% purity.

Conclusions:

Synergistic enzyme engineering and process intensification successfully boosted lactulose biosynthesis efficiency.
Developed viable enzyme variants and system solutions for industrial-scale lactulose production.
Structural insights revealed mechanisms for improved enzyme performance, including enhanced rigidity and allosteric regulation.