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CN-122011135-A - Mitochondrion targeting signal peptide, and coding gene and application thereof

CN122011135ACN 122011135 ACN122011135 ACN 122011135ACN-122011135-A

Abstract

The invention discloses a mitochondrion targeting signal peptide, and a coding gene and application thereof, and belongs to the technical field of biomedicine. The amino acid sequence of the mitochondrion targeting signal peptide is shown as SEQ ID NO.2 or SEQ ID NO. 4. The signal peptide is derived from deep analysis of a Mitochondrial Localization Signal (MLS) region of PCV2 Cap protein, and through accurate confirmation of a 29 th-40 th key amino acid region, the defects of long sequence, unknown structure, unstable targeting efficiency and hidden danger of off-target of the existing natural signal peptide are effectively avoided. The invention not only provides an efficient and safe visualization and positioning means for basic research of mitochondria, but also provides a key technical platform with great potential for developing gene therapy, protein delivery, pharmaceutical preparations and other transformation medicine applications of targeted mitochondria.

Inventors

  • YU WANTING
  • DING ZHEN
  • JIANG QINGQING
  • YANG LIQIN
  • LIU ZEPENG
  • Nie Ruping

Assignees

  • 江西农业大学

Dates

Publication Date
20260512
Application Date
20260212

Claims (10)

  1. 1. The mitochondrial targeting signal peptide is characterized in that the amino acid sequence of the mitochondrial targeting signal peptide is shown as SEQ ID NO.2 or SEQ ID NO. 4.
  2. 2. A nucleic acid molecule encoding the mitochondrial targeting signal peptide of claim 1.
  3. 3. The nucleic acid molecule of claim 2, wherein the nucleotide sequence is set forth in SEQ ID No.8 or SEQ ID No. 10.
  4. 4. A fusion protein comprising the mitochondrial targeting signal peptide of claim 1, and a protein of interest linked to the C-terminus thereof.
  5. 5. The fusion protein of claim 4, wherein the protein of interest comprises a fluorescent protein, an enzyme, a therapeutic protein, an antigenic protein, and a reporter protein.
  6. 6. A recombinant vector comprising the nucleic acid molecule of claim 2 or 3 or a nucleic acid molecule encoding the fusion protein of claim 4 or 5.
  7. 7. Use of a mitochondrial targeting signal peptide according to claim 1, a nucleic acid molecule according to claim 2 or 3, a fusion protein according to claim 4 or 5 or a recombinant vector according to claim 6 for the preparation of a mitochondrial targeting drug delivery system.
  8. 8. Use of a mitochondrial targeting signal peptide according to claim 1, a nucleic acid molecule according to claim 2 or 3, a fusion protein according to claim 4 or 5 or a recombinant vector according to claim 6 for the preparation of a medicament for the treatment of a mitochondrial related disease.
  9. 9. Use of a mitochondrial targeting signal peptide according to claim 1, a nucleic acid molecule according to claim 2 or 3, a fusion protein according to claim 4 or 5 or a recombinant vector according to claim 6 for the preparation of a product for locating mitochondria or indicating mitochondrial position.
  10. 10. Use of a mitochondrial targeting signal peptide according to claim 1, a nucleic acid molecule according to claim 2 or 3, a fusion protein according to claim 4 or 5 or a recombinant vector according to claim 6 for the preparation of a product for studying the interaction of a protein of interest with mitochondria.

Description

Mitochondrion targeting signal peptide, and coding gene and application thereof Technical Field The invention relates to the technical field of biomedicine, in particular to a mitochondrion targeting signal peptide, and a coding gene and application thereof. Background Mitochondria serve as energy metabolism centers of eukaryotic cells, and their function implementation is highly dependent on the precise localization and transport of mitochondrial proteins encoded by nuclear genes. It is currently known that most mitochondrial proteins, when synthesized in the cytoplasm, carry a specific sequence at their N-terminus called the mitochondrial localization signal peptide. The signal peptide can be recognized by a receptor on a mitochondrial membrane, guides a protein precursor to cross the mitochondrial membrane, is excised by a specific enzyme after entering a mitochondrial matrix, and finally completes the maturation and positioning of the protein. This process involves unfolding and refolding of the precursor protein, the assistance of molecular chaperones, and sorting information carried by the different fragments in the signal peptide, which together determine the final subdomain of the protein in the mitochondria. Therefore, the mitochondria localization signal peptide with high efficiency and specificity is a key molecular tool for realizing targeted delivery of exogenous proteins or therapeutic molecules to mitochondria. However, there are certain limitations to the use of existing native mitochondrial localization signal peptides. First, the targeting efficiency and specificity of signal peptides from different sources are different, and partial signal peptides may have unstable localization effect or off-target phenomenon due to different cell types, physiological states or fusion protein properties. Second, the natural signal peptide sequence is often long, may have unpredictable effects on the structure and function of the protein of interest to which it is fused, or may introduce additional risk of immunogenicity or cytotoxicity due to its own containing unnecessary domains. More importantly, there is currently no systematic, deep resolution of the critical amino acid sequences or domains in the signal peptide that determine its targeting function, which makes rational design and optimization of the signal peptide difficult for specific application scenarios (e.g., high-precision organelle tracking, high-efficiency therapeutic molecule delivery). How to identify core functional sequences from known or newly discovered signal peptides and develop on this basis a novel, shorter, more efficient, more stable mitochondrial targeting tool is a major challenge facing current technology. Furthermore, in the field of therapeutic research for mitochondrial related diseases (such as mitochondrial encephalomyopathy), while gene therapy, protein replacement therapy or drug delivery strategies targeting mitochondria have shown great potential, their development is severely limited by the lack of efficient, safe delivery vehicles. The existing delivery systems often have difficulty achieving specific accumulation of therapeutic ingredients within the wire granules, resulting in limited efficacy or off-target side effects. Therefore, the development of a novel mitochondrial targeting signal peptide with clear core functional domain, concise sequence, high targeting efficiency and good biocompatibility has urgent demands and important significance for promoting innovation of mitochondrial biology basic research tools and breaking through the technical bottleneck of targeted treatment of mitochondrial diseases. Disclosure of Invention The invention aims to provide a mitochondria targeting signal peptide, and a coding gene and application thereof, so as to solve the problems in the prior art. The invention provides a novel mitochondrial targeting signal peptide which has a short structure and definite targeting, and overcomes the defects of redundancy and instability of a natural signal peptide by determining 29-40 key functional domains. The fusion protein system constructed based on the signal peptide has high targeting efficiency and good biocompatibility, and provides a high-efficiency and reliable technical platform for mitochondrial research and disease targeting treatment. In order to achieve the above object, the present invention provides the following solutions: the invention provides a mitochondrion targeting signal peptide, the amino acid sequence of which is shown as SEQ ID NO.2 or SEQ ID NO. 4. The invention also provides a nucleic acid molecule encoding the mitochondrial targeting signal peptide. Alternatively, the nucleotide sequence is shown as SEQ ID NO.8 or SEQ ID NO. 10. The invention also provides a fusion protein, which comprises the mitochondrion targeting signal peptide and a target protein connected to the C end of the fusion protein. Alternatively, the protein of interest includes fluoresce