{"entity": "researcher", "timestamp": "2026-08-20T20:54:57.285Z", "family": "Hendrikse", "given": "Natalie M", "initials": "NM", "orcid": "0000-0001-8644-3408", "affiliations": ["School of Engineering Sciences in Chemistry, Biotechnology and Health, Science for Life Laboratory, KTH Royal Institute of Technology, 114 28 Stockholm, Sweden.", "School of Engineering Sciences in Chemistry, Biotechnology and Health, Department of Fibre and Polymer Technology, KTH Royal Institute of Technology, 114 28 Stockholm, Sweden.", "Swedish Orphan Biovitrum AB, 112 76 Stockholm, Sweden."], "links": {"self": {"href": "https://publications-affiliated.scilifelab.se/researcher/711d4343f4594d87852869666b347f98.json"}, "display": {"href": "https://publications-affiliated.scilifelab.se/researcher/711d4343f4594d87852869666b347f98"}}, "publications": [{"entity": "publication", "iuid": "655947c203b44a9f8a690e2cf9961d78", "links": {"self": {"href": "https://publications-affiliated.scilifelab.se/publication/655947c203b44a9f8a690e2cf9961d78.json"}, "display": {"href": "https://publications-affiliated.scilifelab.se/publication/655947c203b44a9f8a690e2cf9961d78"}}, "title": "Engineering of Ancestors as a Tool to Elucidate Structure, Mechanism, and Specificity of Extant Terpene Cyclase.", "authors": [{"family": "Schriever", "given": "Karen", "initials": "K", "orcid": "0000-0002-3677-5508", "researcher": {"href": "https://publications-affiliated.scilifelab.se/researcher/282cce0f6c054fd8a7858977be549b24.json"}}, {"family": "Saenz-Mendez", "given": "Patricia", "initials": "P", "orcid": "0000-0002-6711-4972", "researcher": {"href": "https://publications-affiliated.scilifelab.se/researcher/0efb9bc5eb7344bea1f44272fa4b2540.json"}}, {"family": "Rudraraju", "given": "Reshma Srilakshmi", "initials": "RS"}, {"family": "Hendrikse", "given": "Natalie M", "initials": "NM", "orcid": "0000-0001-8644-3408", "researcher": {"href": "https://publications-affiliated.scilifelab.se/researcher/711d4343f4594d87852869666b347f98.json"}}, {"family": "Hudson", "given": "Elton P", "initials": "EP"}, {"family": "Biundo", "given": "Antonino", "initials": "A"}, {"family": "Schnell", "given": "Robert", "initials": "R"}, {"family": "Syr\u00e9n", "given": "Per-Olof", "initials": "PO", "orcid": "0000-0002-4066-2776", "researcher": {"href": "https://publications-affiliated.scilifelab.se/researcher/b035f26bd71e42f9942f9a4943ed0d1a.json"}}], "type": "journal article", "published": "2021-03-17", "journal": {"title": "Journal of the American Chemical Society", "issn": "1520-5126", "volume": "143", "issue": "10", "pages": "3794-3807", "issn-l": "0002-7863"}, "abstract": "Structural information is crucial for understanding catalytic mechanisms and to guide enzyme engineering efforts of biocatalysts, such as terpene cyclases. However, low sequence similarity can impede homology modeling, and inherent protein instability presents challenges for structural studies. We hypothesized that X-ray crystallography of engineered thermostable ancestral enzymes can enable access to reliable homology models of extant biocatalysts. We have applied this concept in concert with molecular modeling and enzymatic assays to understand the structure activity relationship of spiroviolene synthase, a class I terpene cyclase, aiming to engineer its specificity. Engineering a surface patch in the reconstructed ancestor afforded a template structure for generation of a high-confidence homology model of the extant enzyme. On the basis of structural considerations, we designed and crystallized ancestral variants with single residue exchanges that exhibited tailored substrate specificity and preserved thermostability. We show how the two single amino acid alterations identified in the ancestral scaffold can be transferred to the extant enzyme, conferring a specificity switch that impacts the extant enzyme's specificity for formation of the diterpene spiroviolene over formation of sesquiterpenes hedycaryol and farnesol by up to 25-fold. This study emphasizes the value of ancestral sequence reconstruction combined with enzyme engineering as a versatile tool in chemical biology.", "doi": "10.1021/jacs.0c10214", "pmid": "33496585", "labels": [], "xrefs": [{"db": "pmc", "key": "PMC8023661"}], "notes": [], "created": "2026-08-20T08:45:04.766Z", "modified": "2026-08-20T08:45:04.843Z"}, {"entity": "publication", "iuid": "627b5913195446589f73aeaf2cc1ab7d", "links": {"self": {"href": "https://publications-affiliated.scilifelab.se/publication/627b5913195446589f73aeaf2cc1ab7d.json"}, "display": {"href": "https://publications-affiliated.scilifelab.se/publication/627b5913195446589f73aeaf2cc1ab7d"}}, "title": "Exploring the therapeutic potential of modern and ancestral phenylalanine/tyrosine ammonia-lyases as supplementary treatment of hereditary tyrosinemia.", "authors": [{"family": "Hendrikse", "given": "Natalie M", "initials": "NM", "orcid": "0000-0001-8644-3408", "researcher": {"href": "https://publications-affiliated.scilifelab.se/researcher/711d4343f4594d87852869666b347f98.json"}}, {"family": "Holmberg Larsson", "given": "Albin", "initials": "A"}, {"family": "Svensson Gelius", "given": "Stefan", "initials": "S"}, {"family": "Kuprin", "given": "Sergei", "initials": "S"}, {"family": "Nordling", "given": "Erik", "initials": "E"}, {"family": "Syr\u00e9n", "given": "Per-Olof", "initials": "PO"}], "type": "journal article", "published": "2020-01-28", "journal": {"title": "Sci Rep", "issn": "2045-2322", "volume": "10", "issue": "1", "pages": "1315", "issn-l": "2045-2322"}, "abstract": "Phenylalanine/tyrosine ammonia-lyases (PAL/TALs) have been approved by the FDA for treatment of phenylketonuria and may harbour potential for complementary treatment of hereditary tyrosinemia Type I. Herein, we explore ancestral sequence reconstruction as an enzyme engineering tool to enhance the therapeutic potential of PAL/TALs. We reconstructed putative ancestors from fungi and compared their catalytic activity and stability to two modern fungal PAL/TALs. Surprisingly, most putative ancestors could be expressed as functional tetramers in Escherichia coli and thus retained their ability to oligomerize. All ancestral enzymes displayed increased thermostability compared to both modern enzymes, however, the increase in thermostability was accompanied by a loss in catalytic turnover. One reconstructed ancestral enzyme in particular could be interesting for further drug development, as its ratio of specific activities is more favourable towards tyrosine and it is more thermostable than both modern enzymes. Moreover, long-term stability assessment showed that this variant retained substantially more activity after prolonged incubation at 25 \u00b0C and 37 \u00b0C, as well as an increased resistance to incubation at 60 \u00b0C. Both of these factors are indicative of an extended shelf-life of biopharmaceuticals. We believe that ancestral sequence reconstruction has potential for enhancing the properties of enzyme therapeutics, especially with respect to stability. This work further illustrates that resurrection of putative ancestral oligomeric proteins is feasible and provides insight into the extent of conservation of a functional oligomerization surface area from ancestor to modern enzyme.", "doi": "10.1038/s41598-020-57913-y", "pmid": "31992763", "labels": [], "xrefs": [{"db": "pmc", "key": "PMC6987202"}, {"db": "pii", "key": "10.1038/s41598-020-57913-y"}], "notes": [], "created": "2026-08-20T09:04:49.823Z", "modified": "2026-08-20T09:04:49.896Z"}]}