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The fluorination of amino acid residues represents a near-isosteric alteration with the potential to report on biological pathways, yet the site-directed editing of carbon-hydrogen (C-H) bonds in complex biomolecules to carbon-fluorine (C-F) bonds is challenging, resulting in its limited exploitation. Here, we describe a protocol for the posttranslational and site-directed alteration of native γCH 2 to γCF 2 in protein sidechains. This alteration allows the installation of difluorinated sidechain analogs of proteinogenic amino acids, in both native and modified states. This chemical editing is robust, mild, fast and highly efficient, exploiting photochemical- and radical-mediated C-C bonds grafted onto easy-to-access cysteine-derived dehydroalanine-containing proteins as starting materials. The heteroaryl-sulfonyl reagent required for generating the key carbon-centered C• radicals that install the sidechain can be synthesized in two to six steps from commercially available precursors. This workflow allows the nonexpert to create fluorinated proteins within 24 h, starting from a corresponding purified cysteine-containing protein precursor, without the need for bespoke biological systems. As an example, we readily introduce three γCF 2 -containing methionines in all three progressive oxidation states (sulfide, sulfoxide and sulfone) as D-/L- forms into histone eH3.1 at site 4 (a relevant lysine to methionine oncomutation site), and each can be detected by 19 F-nuclear magnetic resonance of the γCF 2 group, as well as the two diastereomers of the sulfoxide, even when found in a complex protein mixture of all three. The site-directed editing of C-H→C-F enables the use of γCF 2 as a highly sensitive, 'zero-size-zero-background' label in protein sidechains, which may be used to probe biological phenomena, protein structures and/or protein-ligand interactions by 19 F-based detection methods., (© 2023. Springer Nature Limited.)

Many pathogens exploit host cell-surface glycans. However, precise analyses of glycan ligands binding with heavily modified pathogen proteins can be confounded by overlapping sugar signals and/or compounded with known experimental constraints. Universal saturation transfer analysis (uSTA) builds on existing nuclear magnetic resonance spectroscopy to provide an automated workflow for quantitating protein-ligand interactions. uSTA reveals that early-pandemic, B-origin-lineage severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) spike trimer binds sialoside sugars in an "end-on" manner. uSTA-guided modeling and a high-resolution cryo-electron microscopy structure implicate the spike N-terminal domain (NTD) and confirm end-on binding. This finding rationalizes the effect of NTD mutations that abolish sugar binding in SARS-CoV-2 variants of concern. Together with genetic variance analyses in early pandemic patient cohorts, this binding implicates a sialylated polylactosamine motif found on tetraantennary N-linked glycoproteins deep in the human lung as potentially relevant to virulence and/or zoonosis.

Objective: To describe the epidemiological features of a possible disease cluster of vulvar cancer and pre-cancers in Australian Indigenous women living in the Northern Territory (NT) of Australia., Methods: We identified NT-resident women with a confirmed histological diagnosis of vulvar cancer or high-grade vulvar intraepithelial neoplasia (VIN) between 1 January 1996 and 31 December 2005., Results: Seventy-one women were identified; 32 diagnosed with vulvar cancer and 39 with high-grade VIN. Most women diagnosed were Indigenous, aged less than 50 years and living in remote communities in the East Arnhem (EA) district, on the north-east coast of the NT. The age-adjusted incidence rate of vulvar cancer in EA Indigenous women aged 0-49 years was 31.1 per 100,000 (95% CI 13.1-49.1), over 50 times higher than the national Australian rate (0.4 per 100,000, 95% CI 0.4-0.5) for the same age-group. In the age-group of 0-49 years, the age-adjusted incidence rate of VIN for EA Indigenous women was 34.7 per 100,000 (95% CI 15.2-54.3), compared with 6.7 per 100,000 (95% CI 2.0-11.4) for Indigenous women living elsewhere in the Top End of the NT., Conclusion: These data provide evidence of a geographic cluster of vulvar cancer in remote Indigenous communities in northern Australia.

Objective: To analyze patterns and frequency of recurrences of squamous cell carcinoma (SCC) of the vulva after wide local excision (WLE) and superficial inguinal lymphadenectomy with separate incisions and to identify prognostic factors for the development of recurrences., Methods: Between January 1985 and December 1999, all 125 consecutive patients with primary SCC of the vulva, treated with WLE and superficial inguinal lymphadenectomy, were retrospectively analyzed. Recurrences were registered by localization as: local, skin bridge, groin or distant., Results: A local recurrence was diagnosed in 29 (23%) patients, 11 (9%) developed a groin and 4 (3%) a distant recurrence. No skin bridge recurrences were identified. The 5 years local relapse-free survival was 70%. After a first local recurrence, 72% of these patients developed a second local recurrence. Adjusted for other predictors, older age (>74 years) is an independent risk factor for local recurrences (HR: 2.38; 95%-C.I.: 1.08-5.23) and stage III/IV cancer for developing groin/distant recurrences (HR: 3.03; 95%-C.I.: 1.0-9.18)., Conclusion: WLE and superficial inguinal lymphadenectomy with separate incisions result in a high groin recurrence rate in this study; superficial lymphadenectomy should be replaced by deep inguinofemoral lymphadenectomy. After a local recurrence, 72% of the patients developed a second local recurrence. These patients are at high risk and need a close follow-up.