Normal and Pathological Red Cell Physiology

A fixed time possition "Engineer in membrane proteins expression in mammal’s cells (18 months)" is open in our laboratory.

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Engineer in membrane proteins expression in mammal’s cells (18 months)

Workplace: Inserm U1134, Integrated Biology of Red Blood Cell (BIGR) 149, rue de Sèvres Paris 15th and Inserm U1135 Centre d’Immunologie et Maladies Infectieuses (CIMI) 91, Boulevard de l’Hôpital Paris 13th .

Type of contract: Fixed Term contract (CDD)

Duration of the contract: 18 months

Expected date of employment: July 15th, 2024

Deadline for application: June 20th, 2024

Applications should be sent to: Mariano Ostuni (mariano.ostuni@inserm.fr) . Don’t hesitate to contact-us if you need more information.

Remuneration: ~ 2100 €/ month (after deduction of payroll taxes and before deduction of the income tax and depending on the experience level)

Minimal Requested Education level: Bac +5 (Engineer, MSc or equivalent)

Requested Experience: 1-2 years

Missions

The engineer will produce erythrocyte membrane proteins (production and purification), which will be thereafter reconstituted into Giant Unilamellar Vesicles (GUVs) by project’s partners. This involves in particular the extraction and purification of membrane proteins under detergent-free conditions, and its inclusion in mimetic membrane systems (SMALPs or nanodiscs/liposomes reconstituted). Proteins should be purified from erythrocyte "ghosts" and/or eukaryotic cell lines expressing recombinant tagged proteins.

Activities

- Construction of expression plasmids for tagged membrane proteins subcloning.

- Transfections and production of recombinant proteins in eukaryotic cells

- Solubilization of membrane proteins

- Proteins’ purification by size exclusion and affinity chromatography methods (Akta or Gravity Flow Columns)

- Biophysical and biochemical characterization of produced proteins

Skills

- Molecular biology and cloning: plasmid constructions, bacterial transformations, mini and maxi plasmid preparations (required)

-Cell biology techniques: eukaryotic cell cultures (required), flow cytometry (suitable), confocal microscopy (suitable).

- Production of recombinant proteins in eukaryotic systems (required)

- Purification of proteins by chromatographic methods on Akta system (suitable)

- Assay and characterization of protein purity (required)

- Solubilization and extraction of membrane proteins in nano-objects (SMA), reconstitutions in GUVs (notions, internal training possible)

Work context

BIGR -Integrated Biology of the Red Blood Cell- is a Research Laboratory from the Inserm and Université Paris Cité, focusing on the properties of human red blood cells (erythrocytes) and located in the 15th district of Paris (www.u1134.inserm.fr).

Our laboratory is mainly interested to determine the gene sequence up to the protein structure of blood group antigens, the molecular analysis of their polymorphisms and the understanding of the mechanisms involved in certain hereditary disorders of the erythroid membrane.

Experimental and in-silico methods are used following an integrative approach to understand the interaction between extracellular ligands, the cytoskeleton and transmembrane proteins which define the antigenic properties (e.g., blood type) of erythrocytes.

These interactions are studied in normal and pathological situations (hereditary anemia, malaria, sickle cell disease) with particular interest in the structure and function of the erythrocyte membrane.

The candidate will join the group "Transporters and complexes of the erythrocyte membrane" led by Mr Ostuni within the Team 1 "Physiology of the Normal and Pathological Red Blood Cell" (https://sites.google.com/view/umrs1134-team1), to develop the SynEry project (Synthetic Erythrocyte) in the framework of a European Consortium including teams from Belgium, Spain and Italy.

This position requires the candidate to have good communication skills, especially in English

2) A Fixed-term Technician position (from march/April 2024 to August 2025) is opened in our team.

POSITION SUMMARY

1) Implement, within the framework of established protocols, biochemistry, cellular and molecular biology techniques for team 1 “Normal and Pathological Red Blood Cell Physiology” under the direction of two researchers from the team.

2) Provide technical support for the hemorheology technical platform under the direction of an engineer and the management of the team and the Unit

PRESENTATION OF THE MANAGEMENT/WELCOMING STRUCTURE OF THE POSITION

Laboratory address: Necker Hospital, Lavoisier Building, 149, rue de Sèvres, 75015, Paris, France

UMR_S1134 BIGR (Integrated Biology of the Red Blood Cell) is under the supervision of Paris Cité University (attached to the UFR SdV, Faculty of Sciences) and Inserm. The UMR is directed by Caroline Le Van Kim and is organized into 4 teams comprising 80 people including 52 permanent staff (18 Teacher-Researchers and UPCité University Hospital, 10 Inserm, CNRS and EFS Researchers, 10 Biatss/ITA and 3 Hospital Practitioners) as well as 35 non-permanent (18 fixed-term contracts and 17 doctoral students). We are developing in cellulo, in vitro and in silico approaches to study the biology of red blood cells (RBCs), from their genesis to their elimination, as well as their interaction with blood cells, vascular compartments and the spleen. We are strongly interested in the mechanisms involved in the pathophysiology of RBC and erythropoiesis diseases, particularly sickle cell disease, a public health priority. Our projects are also focused on transfusion biology and blood groups, an area where we have very high international visibility.

ACTIVITIES

Learn and conduct experiments in the fields of biology (cell culture of immortalized and primary cells and flow cytometry mainly, basic molecular biology such as PCR). Gather and format the results of experiments. Write and update technical protocols. Keep a laboratory notebook. Prepare the equipment and carry out systematic checks and adjustments. Apply the regulations of the field of study. Monitor the equipment, manage the usage schedule and ensure first-level maintenance. Manage stocks and orders. Train in the longer term, and implement rheology experiments linked to the use of the hemo-rheology platform.

The agent will be supervised by two researchers (Mariano Ostuni and Slim Azouzi) from team 1 of the Unit. The hierarchical superiors will be the two researchers and the director of the research team

REQUIRED PROFILE

Knowledge: Basic molecular and cellular biology techniques (cell culture, transfections, PCR, flow cytometry, …), theoretical knowledge in molecular and cellular biology.

Know-how - Operational skills: Know how to implement cell biology, molecular biology and biochemistry techniques. Use software specific to the activity. Know how to write reports. Adapt experimental protocols to specific needs. Keep a laboratory notebook. Communicate your results orally. Be able to place orders using appropriate software.

Soft skills – Behavioral skills: The agent must be able to demonstrate interpersonal skills, organizational skills, critical thinking and intellectual curiosity. He must like working in a team. While being supervised, the agent must have a capacity for autonomy in experiments and experience reports..

Requested Education level: Bac+3 License, Professional License

Application procedures: To submit your application, send your CV and cover letter by email with the reference of the offer to caroline.le-van-kim@inserm.fr ; mariano.ostuni@inserm.fr and to drh.recrutement@u-paris.fr