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To apply this class of nanomaterials in biomedical fields, a thorough understanding of their interaction with biomacromolecules is essential. Given the therapeutic importance of bromelain (BM) proteins, here, the impact of GO concentration and reduction on their interaction with bromelain has been examined using a variety of spectroscopic and biophysical approaches. The results indicate that the adsorption of BM proteins on the surface of GO\/RGO occurs spontaneously and rapidly, forming a protein corona (PC). The binding affinity of BM with GO is much higher than with its reduced form (RGO). The binding of BM with GO occurs through cooperative interactions, whereas both cooperative and anticooperative binding are observed in the BM\/RGO system. The adsorption of BM on GO nanosheets is primarily driven by strong electrostatic and H-bonding interactions, whereas weak hydrophobic interactions are the primary driving forces behind BM adsorption on the RGO surface. Distinct kinds of corona were observed in the GO\/RGO-BM system based on the types of interactions involved (such as electrostatic, hydrophobic, H-bonding) between the protein and nanoparticles. BM exhibited a stronger and more stable association with GO, showing hard-corona-like characteristics, whereas its association with RGO was comparatively weaker and less stable, showing soft-corona-like characteristics. The therapeutic utility of this protein\u2013nanoparticle system will be heavily dependent on its PC formation, which will be the focus of future research. This study lays the groundwork for future PC investigations on the diverse range of protein nanoparticles utilized in nanomedicine and environmental applications.<\/jats:p>","DOI":"10.1021\/acs.jpcb.6c03763","type":"journal-article","created":{"date-parts":[[2026,10,10]],"date-time":"2026-10-10T03:02:23Z","timestamp":1791601343000},"page":"11317-11332","source":"Crossref","is-referenced-by-count":0,"title":["Deciphering the\nRole of the Concentration and Reduction\nof Graphene Oxide Nanosheets in Their Interaction and Protein Corona\n(PC) Formation with Bromelain Proteins: A Spectroscopic and Biophysical\nApproach"],"prefix":"10.1021","volume":"130","author":[{"given":"Raju","family":"Sardar","sequence":"first","affiliation":[{"name":"Jadavpur University , , ,","place":["Kolkata, India, 700032"]}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Himal","family":"Das","sequence":"additional","affiliation":[{"name":"Jadavpur University , , ,","place":["Kolkata, India, 700032"]}],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0000-0001-7628-5768","authenticated-orcid":true,"given":"Soumen","family":"Ghosh","sequence":"additional","affiliation":[{"name":"Jadavpur University , , ,","place":["Kolkata, India, 700032"]}],"role":[{"vocabulary":"crossref","role":"author"}]}],"member":"316","published-online":{"date-parts":[[2026,10,9]]},"reference":[{"key":"2026102207421611700_cit1","doi-asserted-by":"publisher","first-page":"5019","DOI":"10.1021\/nn1006539","article-title":"Hierarchical\nnanocomposites of polyaniline nanowire arrays on graphene oxide sheets\nwith synergistic effect for energy storage","volume":"4","author":"Xu","year":"2010","journal-title":"ACS Nano"},{"key":"2026102207421611700_cit2","doi-asserted-by":"publisher","first-page":"17268","DOI":"10.1021\/am505308f","article-title":"Covalent functionalization\nof graphene oxide with biocompatible poly\n(ethylene glycol) for delivery of paclitaxel","volume":"6","author":"Xu","year":"2014","journal-title":"ACS Appl. 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Acta"},{"key":"2026102207421611700_cit14","doi-asserted-by":"publisher","first-page":"6094","DOI":"10.1039\/C5CS00217F","article-title":"The nanoparticle\nbiomolecule corona: lessons learned\u2013challenge\naccepted?","volume":"44","author":"Docter","year":"2015","journal-title":"Chem. Soc. Rev."},{"issue":"1","key":"2026102207421611700_cit15","doi-asserted-by":"publisher","first-page":"2026","DOI":"10.1002\/EXP.20240248","article-title":"Surface Charge-Determined\nProtein Coronas of Nanoparticles Control\nEndothelial Cells Uptake Under Low Magnitude Shear Stress","volume":"6","author":"Zhang","year":"2026","journal-title":"Exploration"},{"key":"2026102207421611700_cit16","doi-asserted-by":"publisher","first-page":"301","DOI":"10.1039\/C3MH00106G","article-title":"Protein corona formation around nanoparticles \u2013\nfrom the past to the future","volume":"1","author":"Del Pino","year":"2014","journal-title":"Mater. 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Nanobiotechnol."},{"key":"2026102207421611700_cit19","doi-asserted-by":"publisher","first-page":"8978","DOI":"10.1039\/C5NR00520E","article-title":"Personalized disease-specific\nprotein\ncorona influences the therapeutic impact of graphene oxide","volume":"7","author":"Hajipour","year":"2015","journal-title":"Nanoscale"},{"key":"2026102207421611700_cit20","doi-asserted-by":"publisher","first-page":"5761","DOI":"10.1021\/ja910675v","article-title":"What the\ncell \u201csees\u201d in bionanoscience","volume":"132","author":"Walczyk","year":"2010","journal-title":"J. Am. Chem. Soc."},{"key":"2026102207421611700_cit21","doi-asserted-by":"publisher","first-page":"110073","DOI":"10.1016\/j.ecoenv.2019.110073","article-title":"Study on potential\ntoxic material base and mechanisms of hepatotoxicity induced by Dysosma\nversipellis based on toxicological evidence chain (TEC) concept","volume":"190","author":"Liu","year":"2020","journal-title":"Ecotoxicol. Environ. 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Technol."},{"issue":"1","key":"2026102207421611700_cit32","doi-asserted-by":"publisher","first-page":"1","DOI":"10.1002\/imt2.70111","article-title":"Curcuminoids\namplify host innate antiviral immunity via the CRYAB-RBM26 axis in\nviral infection","volume":"5","author":"Chen","year":"2026","journal-title":"iMeta"},{"key":"2026102207421611700_cit33","doi-asserted-by":"publisher","first-page":"154","DOI":"10.1002\/smll.201101695","article-title":"Assembly of graphene\noxide\u2013enzyme conjugates through hydrophobic\ninteraction","volume":"8","author":"Zhang","year":"2012","journal-title":"Small"},{"issue":"1","key":"2026102207421611700_cit34","doi-asserted-by":"publisher","first-page":"195","DOI":"10.1038\/s41522-025-00830-7","article-title":"TNF-\u03b1 modulation by rice bran peptides: implications\nfor gut microbiota stability and cognitive health in aging","volume":"11","author":"Mu","year":"2025","journal-title":"Npj Biofilms Microbiomes."},{"key":"2026102207421611700_cit35","doi-asserted-by":"publisher","first-page":"1975","DOI":"10.1002\/etc.3055","article-title":"Transformation\nof graphene\noxide by ferrous iron: Environmental implications","volume":"34","author":"Wang","year":"2015","journal-title":"Environ. Toxicol. Chem."},{"key":"2026102207421611700_cit36","doi-asserted-by":"publisher","first-page":"772","DOI":"10.1038\/nnano.2013.181","article-title":"Rapid formation of plasma\nprotein corona critically affects nanoparticle pathophysiology","volume":"8","author":"Tenzer","year":"2013","journal-title":"Nat. Nanotech."},{"issue":"9\u201310","key":"2026102207421611700_cit37","doi-asserted-by":"publisher","first-page":"1635","DOI":"10.1002\/smll.201201492","article-title":"Biosafety and bioapplication of nanomaterials\nby designing protein\u2013nanoparticle interactions","volume":"9","author":"Yang","year":"2013","journal-title":"Small"},{"issue":"10","key":"2026102207421611700_cit38","doi-asserted-by":"publisher","first-page":"2249","DOI":"10.3390\/foods10102249","article-title":"Bromelain, a group\nof pineapple proteolytic complex enzymes (Ananas comosus) and their\npossible therapeutic and clinical effects. A summary","volume":"10","author":"Varilla","year":"2021","journal-title":"Foods"},{"issue":"1","key":"2026102207421611700_cit39","doi-asserted-by":"publisher","first-page":"1","DOI":"10.1155\/2012\/976203","article-title":"Properties and therapeutic\napplication of bromelain: a review","volume":"2012","author":"Pavan","year":"2012","journal-title":"Biotechnol.\nRes. 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Sci."},{"issue":"6","key":"2026102207421611700_cit42","doi-asserted-by":"publisher","first-page":"469","DOI":"10.1038\/sj.neo.7900196","article-title":"Bromelain reversibly\ninhibits invasive properties of\nglioma cells","volume":"3","author":"Tysnes","year":"2001","journal-title":"Neoplasia"},{"issue":"3","key":"2026102207421611700_cit43","doi-asserted-by":"publisher","first-page":"283","DOI":"10.3892\/br.2016.720","article-title":"Potential role of bromelain in clinical\nand therapeutic\napplications","volume":"5","author":"Rathnavelu","year":"2016","journal-title":"Biomed. Rep."},{"key":"2026102207421611700_cit44","doi-asserted-by":"publisher","first-page":"8900","DOI":"10.1039\/D3CP05361J","article-title":"Exploration of the\nimpact of graphene oxide, acetylenic gemini, and CTAT on the photophysical\nand aggregation properties of dipolar coumarin 153","volume":"26","author":"Sardar","year":"2024","journal-title":"Phys. Chem. Chem. Phys."},{"issue":"2","key":"2026102207421611700_cit45","doi-asserted-by":"publisher","first-page":"142","DOI":"10.1016\/j.saa.2010.03.002","article-title":"Micro-environmental\ninfluences on the fluorescence of tryptophan","volume":"76","author":"Sun","year":"2010","journal-title":"Spectrochim. Acta, Part A"},{"key":"2026102207421611700_cit46","doi-asserted-by":"publisher","first-page":"8168","DOI":"10.1021\/ja042898o","article-title":"Interpretation\nof protein adsorption:\nsurface-induced conformational changes","volume":"127","author":"Roach","year":"2005","journal-title":"J. Am.\nChem. Soc."},{"key":"2026102207421611700_cit47","doi-asserted-by":"publisher","first-page":"419","DOI":"10.1016\/0014-5793(89)81383-3","article-title":"Stem bromelain:\namino acid sequence and implications\nfor weak binding of cystatin","volume":"247","author":"Ritonja","year":"1989","journal-title":"FEBS Lett."},{"key":"2026102207421611700_cit48","doi-asserted-by":"publisher","first-page":"3510","DOI":"10.1002\/slct.201600524","article-title":"Comprehensive\nevaluation of biomolecular interactions\nbetween protein and amino acid based-ionic liquids: A comparable study\nbetween [Bmim][Br] and [Bmim][Gly] ionic liquids","volume":"1","author":"Bisht","year":"2016","journal-title":"ChemistrySelect"},{"key":"2026102207421611700_cit49","doi-asserted-by":"publisher","first-page":"132368","DOI":"10.1016\/j.ijbiomac.2024.132368","article-title":"Addressing the interaction of stem\nbromelain with different anionic surfactants, below, at and above\nthe critical micelle concentration (cmc) in phosphate buffer at pH\n7: Physicochemical, spectroscopic, & molecular docking study","volume":"271","author":"Das","year":"2024","journal-title":"Int. J. Biol. Macromol."},{"key":"2026102207421611700_cit50","doi-asserted-by":"publisher","first-page":"127282","DOI":"10.1016\/j.ijbiomac.2023.127282","article-title":"Investigation\nof the vesicle-to-micelle transition of 11-amino undecanoic\nacid derived sulphonamide and a comprehensive study of its interaction\nwith protein","volume":"253","author":"Dey","year":"2023","journal-title":"Int. J. Biol. Macromol."},{"key":"2026102207421611700_cit51","doi-asserted-by":"publisher","DOI":"10.1007\/978-0-387-46312-4","volume-title":"Principles of fluorescence spectroscopy","author":"Lakowicz","year":"2006"},{"issue":"5","key":"2026102207421611700_cit52","doi-asserted-by":"publisher","first-page":"3702","DOI":"10.1039\/D5CP02915E","article-title":"Electrostatic control\nof protein\u2013vesicle\ninteractions in catanionic long-chain ionic liquid systems: insights\nfrom human serum albumin","volume":"28","author":"Sardar","year":"2026","journal-title":"Phys. Chem. Chem.\nPhys."},{"issue":"1","key":"2026102207421611700_cit53","doi-asserted-by":"publisher","first-page":"365","DOI":"10.1021\/nn9011187","article-title":"F Interaction of gold\nnanoparticles with common human blood proteins","volume":"4","author":"Lacerda","year":"2010","journal-title":"ACS Nano"},{"key":"2026102207421611700_cit54","doi-asserted-by":"publisher","first-page":"3649","DOI":"10.1021\/ol051220q","article-title":"Highly chemoselective addition of amines to epoxides\nin water","volume":"7","author":"Azizi","year":"2005","journal-title":"Org. Lett."},{"issue":"98","key":"2026102207421611700_cit55","doi-asserted-by":"publisher","first-page":"55290","DOI":"10.1039\/C4RA09613D","article-title":"Interaction\nof graphene oxide with human serum albumin\nand its mechanism","volume":"4","author":"Ding","year":"2014","journal-title":"RSC Adv."},{"key":"2026102207421611700_cit56","doi-asserted-by":"publisher","first-page":"8130","DOI":"10.1039\/C7NJ01227F","article-title":"Construing\nthe interactions between MnO2 nanoparticle and bovine serum\nalbumin: insight into the structure and stability of a protein\u2013nanoparticle\ncomplex","volume":"41","author":"Baral","year":"2017","journal-title":"New J. Chem."},{"key":"2026102207421611700_cit57","doi-asserted-by":"publisher","first-page":"10435","DOI":"10.1021\/acs.jpcb.8b08173","article-title":"Exploring\nthe effect of choline-based ionic liquids on the stability and activity\nof stem bromelain","volume":"122","author":"Kumar","year":"2018","journal-title":"J. Phys. Chem. 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