{
  "abstract": "Acid proteases can improve nitrogen availability during ethanol fermentation by hydrolysing feedstock proteins, but their catalytic performance at relatively low temperatures may be limited. In this study, an acid protease-producing Rhodotorula mucilaginosa strain, TSM-6, was isolated from mangrove sediment, and an acid protease gene, rmap9, was identified by whole-genome sequencing and candidate-gene screening. RMAP9 was constitutively expressed in Komagataella phaffii GS115, and secretion was enhanced through EXG1-derived secretion-signal engineering, tandem assembly of three expression cassettes, and fermentation optimisation. Molecular dynamics simulations and Rosetta-based ΔΔG scoring were further used to identify candidate residues for temperature-adaptation engineering, followed by the site-saturation mutagenesis of Ser125. The optimised recombinant strain reached a maximum extracellular acid protease activity of 312.07 ± 18.37 U/mL, representing a 223% increase over the parental constitutive strain. The S125K variant showed a shift in optimum temperature from 50 to 40 °C and approximately 50% higher relative activity at 40 °C than wild-type RMAP9. Moreover, the low-temperature-favoured variants retained more than 80% residual activity after 72 h at 30 °C. These variants combined improved catalytic performance at relatively low temperatures with long-term stability at 30 °C, thereby providing enzyme properties compatible with temperature-related operational requirements for industrial ethanol fermentation.",
  "abstract_shown": true,
  "affiliations": [
    "College of Life Science and Technology, Guangxi University, Nanning 530004, China.",
    "Institute of Biological Science and Technology, Guangxi Academy of Sciences, Nanning 530007, China."
  ],
  "article_type": "research article",
  "arxiv": null,
  "author_total": 7,
  "authors": [
    {
      "affs": [
        0
      ],
      "name": "Yibo Zhang"
    },
    {
      "affs": [
        0
      ],
      "name": "Jiasheng Cao"
    },
    {
      "affs": [
        1
      ],
      "name": "Mingming Luo"
    },
    {
      "affs": [
        0
      ],
      "name": "Zhijie Huang"
    },
    {
      "affs": [
        0
      ],
      "name": "Yu Tang"
    },
    {
      "affs": [
        0
      ],
      "name": "Zhizheng Zhao"
    },
    {
      "affs": [
        0
      ],
      "name": "Jian Lu"
    }
  ],
  "bibtex": "https://research.draconitic.com/paper/3cv9hwqd1b2a0.bib",
  "cited_by": 0,
  "doi": "10.3390/biotech15040087",
  "fulltext_url": "https://europepmc.org/articles/PMC13702498",
  "id": "3cv9hwqd1b2a0",
  "issns": [
    "2673-6284"
  ],
  "landing_url": "https://europepmc.org/article/MED/42930064",
  "language": "eng",
  "licence": "CC-BY",
  "licence_label": "CC BY",
  "licence_source": "article",
  "licence_url": "https://creativecommons.org/licenses/by/4.0/",
  "metadata_licence": null,
  "native_subjects": [
    [
      "openalex",
      "1305|Enzyme Production and Characterization"
    ]
  ],
  "openalex": "W7220654724",
  "pdf_url": "https://europepmc.org/articles/PMC13702498?pdf=render",
  "pmcid": "PMC13702498",
  "pmid": "42930064",
  "published": "2026-10-06",
  "publisher": null,
  "ris": "https://research.draconitic.com/paper/3cv9hwqd1b2a0.ris",
  "sources": [
    {
      "first_seen": "2026-10-30T12:22:01Z",
      "native_id": "MED:42930064",
      "source": "europepmc",
      "url": "https://europepmc.org/article/MED/42930064"
    }
  ],
  "subjects": [
    {
      "field": "biology",
      "origin": "openalex:1305|Enzyme Production and Characterization",
      "subfield": "systems-synthetic"
    }
  ],
  "title": "High-Level Constitutive Secretory Expression and Low-Temperature Adaptation Engineering of the Acid Protease RMAP9 from the Mangrove-Derived Yeast Rhodotorula mucilaginosa.",
  "title_text": "High-Level Constitutive Secretory Expression and Low-Temperature Adaptation Engineering of the Acid Protease RMAP9 from the Mangrove-Derived Yeast Rhodotorula mucilaginosa.",
  "type": "peer-reviewed",
  "updated_at": "2026-10-30T12:22:01Z",
  "url": "https://research.draconitic.com/paper/3cv9hwqd1b2a0",
  "venue": "Biotech (Basel (Switzerland))",
  "versions": []
}