Research

Overview

21/01/2021

Paenarthrobacter nicotinovorans (formerly Arthrobacter nicotinovorans, Arthrobacter oxidans) is a coryneform bacteria able to grow on nicotine as sole carbon and nitrogen source due to the presence of the 165 kb megaplasmid pAO1. The megaplasmid was sequenced and contains 2 major gene clusters: one related to nicotine metabolism and a second cluster forming an operon related to sugar catabolism.





The pAO1 catabolic megaplasmid of Paenarthrobacter nicotinovorans (shades of blue - genes related to nicotine uptake and utilization, green - xylose catabolism related genes, red - recombination related genes, white - unknown function)














I am mainly focused on the molecular biology of the pAO1 megaplasmid related to the defense mechanism against nicotine-generated oxidative stress, sugar-catabolism and biotechnological applications.

Using a unique combination of BLAST searches, protein modeling and in-silico docking I identified putative substrates for the pAO1 coded sugar-operon. The selected compounds were experimentally tested and xylose was identified as the physiological substrate.


The operon coding for a oxidative xylose-catabolic pathway on the pAO1 megaplasmid.


In the last couple of years I focused on the possible medical applications of the nicotine-derivatives produced by Paenarthrobacter nicotinovorans. Using in-silico docking experiments, we tested the potential of nicotine and piridine derivatives produces by this bacteria to bind nAchR and identified 6-Hydroxy-Nicotine (6HNic)  as a potential ligand. In collaboration with  dr. Lucian Hritcu we showed that indeed  6HNic is able to modulate the function of nAChRs and sustain spatial memory formation in a rat model of Alzheimer’s disease. Currently we are working on a biotechnological method to produce and isolate 6HNic using a genetically engineered  Paenarthrobacter nicotinovorans strain.


    


Growing Paenarthrobacter nicotinovorans in a 3L micro-fermenter.






48 hour of growth accelerated in 1 min and 30 secs thanks to PhD Eugen Ungureanu


The latest projects involves the proteomics, transcriptomics and genomic analysis of Paenarthrobacter nicotinovorans pAO1 and its nicotine degradation pathway. Till now, the lab has produced several relevant datasets:

  • the first complete and annotated genome of an nicotine-degrading bacteria Paenarthrobacter nicotinovorans ATCC 49199: GenBank CP089293 and CP089294, as well as the associated raw data: PRJNA693273

  • a transcriptomics data-set dealing with the nicotine metabolism in Paenarthrobacter nicotinovorans ATCC 49199: Gene Expression Omnibus GSE240220

  • a shotgun proteomics dataset that identifies the proteins expressed by Paenarthrobacter nicotinovorans in the presence of nicotine by using nanoLCMSMS PXD008756

  •  another shotgun proteomics dataset that performs a time-based analysis of the nicotine catabolism in Paenarthrobacter nicotinovorans - PXD012577
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Funding

4/04/2021

  • PN-III-P4-ID-PCE-2020-0656, Sequencing the genome of a useful bacteria: Paenarthrobacter nicotinovorans – next step in extending it’s biotechnological applications

  • PN-III-P2-2.1-TE-2016-0367 - Developing an Arthrobacter nicotinovorans biotechnology for neuro-pharmaceuticals production 

  • PN-III-P2-2.1-PED-2016-0177 - Steps towards an Arthrobacter nicotinovorans biotechnology for neuro-pharmaceuticals production

  • PN-II 50BM/2016 - Romania - P.R.P. China joint research project - Nicotine - from toxic residue to metabolic derivatives with neuroprotective effects

  • PN-II-RU-TE-2014-4-0106 – Effects of 6-hydroxy-nicotine on chlorisondamine-induced oxidative stress and neurotoxicity: relevance for Alzheimer’s disease

  • GI-2014-02 – Internal Research grant awarded by the Alexandru I Cuza University of Ia?i

  • TD-236 research grant, Molecular implications of the pAO1 megaplasmid for the Arthrobacter nicotinovorans

  • PD-337 research grant, Cloning and characterization of ORF32 and ORF40 from the pAO1 megaplasmid of Arthrobacter nicotinovorans -potential protein-tagatose interactions models

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Selected publications

Papers/Chapters published in the last 5 years

12/01/2024

  1. Boiangiu, R.S.; Brinza, I.; Honceriu, I.; Mihasan, M.; Hritcu, L. Insights into Pharmacological Activities of Nicotine and 6-Hydroxy-L-nicotine, a Bacterial Nicotine Derivative: A Systematic Review. Biomolecules 2024, 14, 23. [Free FullText]
  2. El-Sabeh, A., Mlesnita, AM., Munteanu, IT. .... Mihasan M. Characterisation of the Paenarthrobacter nicotinovorans ATCC 49919 genome and identification of several strains harbouring a highly syntenic nic-genes cluster. BMC Genomics 24, 536 (2023). [Free Fulltext]
  3. Pérez-Ramos, A.; Ladjouzi, R.; Mihasan, M.; Teiar, R.; Benachour, A.; Drider, D. Advances in Characterizing the Transport Systems of and Resistance to EntDD14, A Leaderless Two-Peptide Bacteriocin with Potent Inhibitory Activity. Int. J. Mol. Sci. 2023, 24, 1517. [Free Fulltext]
  4. Amada El-Sabeh, Iasmina Honceriu, Fakhri Kallabi, Razvan-Stefan Boiangiu, Marius Mihasan - Complete Genome Sequences of Two Closely Related Paenarthrobacter nicotinovorans Strains. Microbiol Resour Announc. 2022; 11(6):e0013322. doi: 10.1128/mra.00133-22. [Fulltext]
  5. Madi-Moussa, D.; Deracinois, B.; Teiar, R.; Li, Y.; Mihasan, M.; Flahaut, C.; Rebuffat, S.; Coucheney, F.; Drider, D. Structure of Lacticaseicin 30 and Its Engineered Variants Revealed an Interplay between the N-Terminal and C-Terminal Regions in the Activity against Gram-Negative Bacteria. Pharmaceutics 2022, 14, 1921.
    doi:10.3390/pharmaceutics14091921 [Fulltext]
  6. Postu, P.A.; Mihasan, M.; Gorgan, D.L.; Sadiki, F.Z.; El Idrissi, M.; Hritcu, L. Pinus halepensis Essential Oil Ameliorates Aβ1-42-Induced Brain Injury by Diminishing Anxiety, Oxidative Stress, and Neuroinflammation in Rats. Biomedicines 2022, 10, 2300. doi:10.3390/biomedicines10092300 [Fulltext]
  7. Mihasan M., Boiangiu R.S., Guzun D., Babii C., Aslebagh R., Channaveerappa D., Dupree D., Darie CC. Time-dependent analysis of Paenarthrobacter nicotinovorans pAO1 nicotine-related proteome. ACS Omega 2021, 6, 22, 14242–14251 [Fulltext]
  8. Mihasan, M. A Beginner’s Guideline for Low?cost 3D Printing of Macromolecules Usable for Teaching and Demonstration. Biochem. Mol. Biol. Educ. 2021, bmb.21493. [Fulltext]
  9. Postu, P. A.; Tiron, A.; Tiron, C. E.; Gorgan, D. L.; Mihasan, M.; Hritcu, L. Conifer Essential Oils Reversed Amyloid Beta1-42 Action by Modulating BDNF and ARC Expression in The Rat Hippocampus. CNS Neurol. Disord. - Drug Targets 2021, 20. [Fulltext]
  10. Boiangiu, R. S.; Mihasan, M.; Gorgan, D. L.; Stache, B. A.; Hritcu, L. Anxiolytic, Promnesic, Anti-Acetylcholinesterase and Antioxidant Effects of Cotinine and 6-Hydroxy-L-Nicotine in Scopolamine-Induced Zebrafish (Danio Rerio) Model of Alzheimer’s Disease. Antioxidants 2021, 10 (2), 212. [Free Fulltext]
  11. Brandsch, R and Mihasan M, A soil bacterial catabolic pathway on the move: Transfer of nicotine catabolic genes between Arthrobacter genus megaplasmids and invasion by mobile elements, Journal of Biosciences, 2020, 45, 1-12  [Fulltext]
  12. Djeussi, D. E.; Noumedem, J. A. K.; Mihasan, M.; Kuiate, J. R.; Kuete, V. Antioxidant Activities of Methanol Extracts of Thirteen Cameroonian Antibacterial Dietary Plants. J. Chem. 2020, 2020. [Free Fulltext]
  13. Dupree EJ, Jayathirtha M, Yorkey H, Mihasan M, Petre BA, Darie CC, A Critical Review of Bottom-Up Proteomics: The Good, the Bad, and the Future of this Field, Proteomes, 2020, 8 (3), 14 [Fulltext]
  14. Boiangiu RS, Mihasan M, Gorgan DL, Stache BA, Petre BA, Hritcu L., Cotinine and 6-Hydroxy-L-Nicotine Reverses Memory Deficits and Reduces Oxidative Stress in A?25-35-Induced Rat Model of Alzheimer’s Disease, Antioxidants, 2020, 9(8), 768 [Fulltext]
  15. Mih??an M, Babii C, Aslebagh R, Channaveerappa D, Dupree EJ, Darie CC. Exploration of Nicotine Metabolism in Paenarthrobacter nicotinovorans pAO1 by Microbial Proteomics. In: Woods AG, Darie CC, editors. Advancements of Mass Spectrometry in Biomedical Research, vol. 1140 of Advances in Experimental Medicine and Biology. 2019. p. 515–29. [Fulltext]
  16. Mih??an M, Wormwood KL, Sokolowska I, Roy U, Woods AG, Darie CC. Mass Spectrometry- and Computational Structural Biology-Based Investigation of Proteins and Peptides. In: Woods AG, Darie CC, editors. Advancements of Mass Spectrometry in Biomedical Research, vol. 1140 of Advances in Experimental Medicine and Biology. 2019. p. 265–87.[Fulltext]

The full list of publications is available here


05/10/2023

  Posters/Presentations in the last 5 years

  1. El-Sabeh, A., Mlesnita, AM., Munteanu, IT. .... Mihasan M., A highly syntenic nic-genes cluster is present in several related bacterial strains, Conferinta Societatii Nationale de Biochimie si Biologie Moleculara, 12-15 Septembrie, Cluj, Romania, [Photo] [Poster].
  2. Marius Mihășan, Abdelrahman Mohamed Taha Mahmoud, Putting figures from Lehninger’s textbook on the table using 3D printing47th FEBS Congress – ‘Together in bioscience for a better future’, Tours, France,  8th - 12th July 2023,  [Poster].
  3. El-Sabeh, A., Mlesnita, AM., Munteanu, IT. .... Mihasan M., A highly syntenic nic-genes cluster is present in several related bacterial strains, 47th FEBS Congress – ‘Together in bioscience for a better future’, Tours, France,  8th - 12th July 2023, [Photo] [Poster].
  4. Mihasan Marius - Full omics caracterisation of a bacteria. Case study - Paenarthrobacter nicotinovorans pAO1 and its nicotine metabolism, Salerno University, Host Assoc. Prof. Anna Maraboti, May 26, 2023 [Presentation]
  5. Mihasan Marius - 3D printed macromolecular models usable for teaching and demonstration, Salerno University, Host Assoc. Prof. Anna Maraboti, May 26, 2023 [Photo] [Photo] [Presentation]
  6. Mihasan Marius - Multi-omics characterisation of nicotine metabolism in Paenarthrobacter nicotinovorans, The 2023 RoBioinfo Conference, 11th-13th May 2023,  [Photo] [Photo] [Presentation]
  7. Mihasan Marius - Steps towards the multi-omics characterization of nicotine metabolism in Paenarthrobacter nicotinovorans ATC 49919, Smart Diaspora 2023, Timisoara, 10th-13th April 2023 [Presentation]
  8. Mihasan Marius - Lessons learned from 3D printing macromolecular models for teaching and demonstration-  The IUBMB-FEBS-PABMB 2022 Congress - The Global Biochemistry Summit, 9-14 July 2022, Lisabona, Portugal, Best Education section poster award by SPB [Photo] [Photo] [Photo] [Photo] [Poster]
  9. I.T. Munteanu, A.M. Mleșnița, F. Kallabi, M. Mihașan - In silico evaluation of antibiotic resistance and CRISPR systems of Paenarthrobater nicotinovorans - The IUBMB-FEBS-PABMB 2022 Congress - The Global Biochemistry Summit, 9-14 July 2022, Lisabona, Portugal*
  10. A. El-Sabeh, I. Honceriu, F. Kallabi, R. Boiangiu, M. Mihașan - Genome sequence of Paenarthrobacter nicotinovorans strain ATCC 49919, a nicotine- degrading soil microorganism, The IUBMB-FEBS-PABMB 2022 Congress - The Global Biochemistry Summit, 9-14 July 2022, Lisabona, Portugal*
  11. Amada El-Sabeh, I. Honceriu, F. Kallabi, R. Boiangiu, M. Mihășan. - Complete genome sequence of the well-studied nicotine-degrading microorganism Paenarthrobacter nicotinovorans strain ATCC 49919, FEMS Conference on Microbiology, June 29 - July 2nd, 2022, Belgrade, Serbia [Photo] [Poster]
  12. Mihasan M., Boiangiu R.S., Guzun D., Babii C., Aslebagh R., Channaveerappa D., Dupree D., Darie CC. - Proteome-wide analysis of Paenarthrobacter nicotinovorans ATCC 49919 nicotine- metabolism, FEMS Conference on Microbiology, June 29 - July 2nd, 2022, Belgrade, Serbia [Photo] [Poster]
  13. Mihasan M., Boiangiu R.S., Guzun D., Babii C., Aslebagh R., Channaveerappa D., Dupree D., Darie CC., Integration and regulation of nicotine metabolism in Paenarthrobacter nicotinovorans as revealed by proteomic data.
    70th ASMS Conference on Mass Spectrometry and Allied Topics, June 5-9 2022, Minneapolis, Minnesota, USA [Photo] [Photo] [Poster]
  14. Mihasan, M. 3D printed macromolecular models usable for teaching and demonstration Marmara University, Host Dr. Beste Turanli June 2022, Istanbul, Turkey [Photo] [Photo]
  15. Mihasan, M. Full omics characterization of a bacteria.  Case study - Paenarthrobacter nicotinovorans pAO1 and its nicotine metabolism,  Marmara University, Host Dr. Beste Turanli June 2022, Istanbul, Turkey [Photo] [Photo]
  16. Mihasan, M. Full omics characterization of a bacteria.  Case study - Paenarthrobacter nicotinovorans pAO1 and its nicotine metabolism,  Clarkson University, Host Dr. Costel C. Darie, June 2022, Potsdam, NY, USA, [Flyer] [Photo] [Presentation]
  17. Mihasan, M., Genome sequencing as a tool to unlock the full biotechnological potential of bacteria,  Clarkson University, School of Arts & Sciences, Walsh Seminar Series, Host Dr. Costel C. Darie, April 2022, Potsdam, NY, USA, Virtual [Flyer] [Presentation]
  18. Mihasan, M. 3D printed macromolecular models usable for teaching and demonstration, November 2020, Lille, France  [Photo] [Photo] [Presentation]
  19. Mihasan, M. A guideline for 3D printing of macromolecular models on the cheap, EMBL Conference: Bringing Molecular Structure to Life: 50 Years of the PDB, 20 - 22 October 2021, Virtual, [Poster]
  20. Mihasan Marius - Mihasan M  - Using molecular models for a better education in Life Sciences, October 2021, Iasi, Romania, [Photo] [Photo] [Presentation]
  21. Mihasan Marius, F. Kallabi - Draft genome of a useful bacteria - Paenarthrobacter nicotinovorans, The 45th FEBS Congress - Molecules of Life: Towards new horizons, 2-8 July, 2021, Virtual,  [Poster]
  22. Mihasan Marius - A guideline for 3D printing of macromolecular models on the cheap, The 45th FEBS Congress - Molecules of Life: Towards new horizons, 2-8 July, 2021, Virtual,  [Poster]
  23. Mihasan, M. A guideline for 3D Printing of macromolecular models on the cheap, PDB50: A special symposium celebrating the 50th anniversary of the Protein Data Bank, May 4, 2021 - May 5, 2021, Virtual [Poster] [Poster presentation]
  24. Mihasan, M., 3D printed macromolecular models usable for teaching and demonstration,  Clarkson University, School of Arts & Sciences, Walsh Seminar Series, Host Dr. Costel C. Darie, April 2021, Potsdam, NY, USA, Virtual [Flyer] [Presentation]
  25. Mihasan M., Brandsch R., Steps towards revealing the evolution of nicotine catabolic gene clusters in Arthrobacters, EMBO conference Molecular Mechanisms in Evolution and Ecology, 30 September - 2nd October 2020, Virtual, [Poster]
  26. Mihasan M, Babii C, Aslebagh R, Channaveerappa D, Dupree EJ, Darie CC. Proteomic Analysis of Nicotine Metabolism in Paenarthrobacter Nicotinovorans, Eastern Analytical Symposium, November 16-19, 2020 [Presentation Abstract][Recorded presentation]
  27. Darie CC, Dupree EJ, Jayathirtha M, Yorkey H, Mihasan M, Petre BA, Proteomics: The Good, the Bad, and the Future of this Field, Eastern Analytical Symposium, November 16-19, 2020 [Presentation Abstract]
  28. Mihasan, M. A surprisingly useful soil microorganism: Paenartharobacter nicotinovorans pAO1,IUT Lille, Host Dr. Francois Krier, December 2019, Lille, France  [Photo] [Photo] [Presentation]
  29. Marius Mihasan, Cornelia Babii, Roshanak Aslebagh, Devika Channaveerappa, Emmalyn Dupree, Costel C. Darie, Proteomics based analysis of the nicotine catabolism in Paenarthrobacter nicotinovorans pAO1, Conferin?a National? a SRBBM, Ia?i, [Photo] [Photo] [Presentation]
  30. M. Mihasan, C. Babii, D. Channaveerappa, R. Aslebagh, E. Dupree, C. Darie - Time-dependent analysis of Paenarthrobacter nicotinovorans pAO1 nicotine-related proteome, 44th FEBS Congress, From Molecules to Living Systems, Krakow, Poland, July 6?11, 2019, poster USAP-39-007 [Abstract] [Photo] [Photo] [Photo] [Poster]
    *Talks and posters presented by coordinated students.

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Colaborations

20/07/2017

  • Dr. Costel C. Darie Biochemistry and Proteomics Group, Department of Chemistry & Biomolecular Science, Clarkson University, Potsdam, NY, USA - kindly accepted me as a Fulbright Fellow for 6 month and provided lab space, reagents, instruments for solving the nicotine induced proteome of Arthrobacter nicotinovorans


Dr. Darie's group after an department seminar, spring 2017. From left to right Dr. Darie, Kelly W., Roshi A., Devika C., Emma D. and Mr. Mihasan wearing the pig T-shirt.  




  •   Dr. Hongzhi Tang (Robert) and  Dr. Fei Tao, Group for Food & Environmental Micro-Biotechnol, State Key Laboratory of Microbial Metabolism, School of Life Sciences & Biotechnology, Shanghai Jiao Tong University


Visiting the School of Life Sciences & Biotechnology, Shanghai Jiao Tong University.
From left to right, Dr. Fei Tao, Hritcu L., Olteanu Z., Stefan M, me, and Dr. Hongzhi Tang on the extreme right.

  • The group of prof. dr. Roderich Brandsch, Institut für Biochemie und Molekularbiologie kindly provided the Arthrobacter nicotinovorans pAO1+ and pAO1- strains, as well as various other E.coli strains, plasmids and reagents. The pAO1 megaplamid was sequenced by this group, so they also provided valuable information regarding the molecular biology of this catabolic plasmid.

Prof. dr. Roderich Brandsch explaining his art concept, September 2010, Freiburg (from left to right: dr. Marius Mihasan, prof. dr. Roderich Brandsch, dr. Eugen Ungureanu) 


  • All enzymes assays, enzyme kinetics and  sugar biochemistry problems are solved with the help of prof. dr. Vlad Artenie, Alexandru I Cuza University, Faculty of Biology 


  • Most of the microbiology related details and practical work involved in this project are closely discussed with associate professor habil. dr. Marius Stefan, Alexandru I Cuza University, Faculty of Biology.



Marius Stefan performing a CASP-3 assay, December 2010, Iasi.

  • Most of the problems dealing with manuscript writing ans submitting, neuroreceptors and transmitters are solved with the help of associate professor habil.  dr. Lucian Hritcu



Last  man standing - dr. Lucian Hritcu the last to leave the Auditorium at the EMBO Meeting 2011, Vienna, Austria


  • All bunch of adminstrative matters, as well as basic chemistry problems are closely discussed with associate professor dr. Zenovia Olteanu

Visiting students
Jaurès AK Noumedem, PhD student, Department of Biochemistry, Faculty of Science, University of Dschang, Dschang, Cameroon, AUF ''Eugen Ionescu'' scholarship, 2011 Julie Reghem, master student, DUT “Genie Biologique”, ABB – Lille, Erasmus Placements, 2013 Victor Baumont, Bsc student, DUT “Genie Biologique”, ABB – Lille, Erasmus Placements, 2015



Fakhri Kallabi, PostDoc, University of SFAX, Tunisia, AUF ''Eugen Ionescu'' scholarship, 2020


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Links

16/12/2014

Funding organisations for romanian universities and postgraduate research programmes:


  • The National Authority for Scientific Research: ANCS

  • Executive Unit for Financing Higher Education, Research, Development and Innovation: UEFISCDI


Protein homology and molecular docking software:

  • Comparative Microbial Genomics BioTools  - CMGBiotools

  • open-source, modular, flexible, molecular modelling and visualization environment - OpenStructure

  • A fully automated protein structure homology-modeling server and model validation  - SWISS-MODEL

  • An alternative way to access and interact with SWISS-MODEL - DeepView (Swiss Pdb-Viewer)

  • Homology or comparative modeling of protein three-dimensional structures, runs locally only via command line (no GUI) - MODELLER

  • A graphical user interface to Modeller program - MINT

  • Automatted homology modelling, flexible protein-protein docking and model validation BISKIT

  • A extensible program for interactive visualization, editing and analysis of molecular structures  - UCSF Chimera

  • Graphical tool for computational protein engineering, mainly site-directed mutagenesis -  TRITON

  • Docking program and tools that must be run localy in a command line DOCK 6

  • Suite of automated docking tools - AutoDock and its free GUI - ADT

  • The new and significantly improved in the average accuracy of the binding mode predictions as well as speed AutoDock Vina

  • One of the most popular programs for molecular visualization - PyMOL with impressive extensibility using various plug-ins:
            performing and visualising docking results with AutoDock/Vina,
            visualization of substrate pathways and entrance tunnels in protein structures CAVER
            visualization of pockets and voids in protein structures detected by CASTp
            create animations and movies with molecular structures - eMovie

  • Bootable, self-configuring system for virtual screening using AutoDock4/Vina - MOLA

Bioinformatics software and Linux:

  • Comparative Microbial Genomics BioTools  - CMGBiotools

  • The swiss army of bioinformatics workstations - Bio-linux

  • A project that aims to develop Debian into an operating system well fit for the requirements for medical practice and biomedical research Debian Med

  • collection of open-source applications for life scientists and an in-house project at Bioinformatics.Org BioBrew

  • A nice commentary on Open software for biologists

Databases and other usefull links:

  • On-line methods collection: Protocol on-line

  • European Bioinformatics Institute: EMBL-EBI

  • Medline at NIH: PubMed

  • Central European Science Publishers: Versita

  • SIB PROTEOMICS: Expasy

  • World Wide Protein Structure Database: wwPDB and its european partner PDBe

  • DNA and protein Sequnce Manipulation Suite: on its original website SMS or mirored by this website SMS2

Life sciences related comercial companies:

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    Contact: marius.mihasan@uaic.ro

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