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Related Experiment Video

Updated: Jul 14, 2026

Expression of Recombinant Proteins in the Methylotrophic Yeast Pichia pastoris

Expression of Recombinant Proteins in the Methylotrophic Yeast Pichia pastoris

Published on: February 26, 2010

Human lysozyme secretion increased by alpha-factor pro-sequence in Pichia pastoris.

C Oka¹, M Tanaka, M Muraki

¹Chiba Prefectural Industrial Research Institute, Chiba, Japan. coka@iri.pref.chiba.jp

Bioscience, Biotechnology, and Biochemistry

|January 15, 2000

Summary

The Saccharomyces alpha factor prepro sequence (MF-alpha Prepro) significantly enhances human lysozyme secretion in Pichia pastoris. This optimized prepro sequence is crucial for high-level, native lysozyme production.

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Published on: January 23, 2019

Area of Science:

Biotechnology
Molecular Biology
Protein Engineering

Background:

High-level secretion of recombinant proteins is essential for industrial applications.
Pichia pastoris is a widely used host for heterologous protein expression.
Optimizing signal sequences is critical for efficient protein secretion.

Purpose of the Study:

To evaluate different signal sequences for high-level human lysozyme secretion in Pichia pastoris.
To determine the role of the Saccharomyces alpha factor prepro sequence (MF-alpha Prepro) in lysozyme secretion.
To engineer leader sequences for enhanced native human lysozyme production.

Main Methods:

Comparative analysis of four signal sequences (chicken lysozyme, leucine-rich artificial, Saccharomyces invertase, MF-alpha Prepro).
Construction and evaluation of mutant MF-alpha Prepro leader sequences.
Characterization of secreted human lysozyme using N-terminal sequencing, mass spectrometry, and crystallography.

Main Results:

MF-alpha Prepro resulted in a 20-fold increase in lysozyme secretion compared to other signal sequences.
Eliminating the pro region of MF-alpha Prepro significantly decreased secretion.
MF-alpha Prepro without spacer sequences yielded native human lysozyme, confirmed by multiple analytical techniques.

Conclusions:

The MF-alpha Prepro sequence is highly effective for high-level secretion of human lysozyme in Pichia pastoris.
The pro region of MF-alpha Prepro plays a vital role in facilitating efficient secretion.
Optimized MF-alpha Prepro constructs enable the production of authentic, non-fused human lysozyme.