Information Package / Course Catalogue - Adnan Menderes University

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Protein Isolation and Identification Techniques

Course Code	: BYK606
Course Type	: Required
Couse Group	: Third Cycle (Doctorate Degree)
Education Language	: Turkish
Work Placement	: N/A

Theory	: 2
Prt.	: 2
Credit	: 3
Lab	: 0
ECTS	: 5

Objectives of the Course

It is aimed to give new developments in the field of protein purification in the light of basic techniques.

Course Content

Structural properties and investigation of proteins, amino acids and chemical proporties, primary structure of proteins: the amino acid sequence, N-terminal and C-terminal, amino acid sequencing / gene relationship, levels of the upper structure of proteins – secondary - tertiary and quaternary structure - the structure - function relationships, proteins and biomembranes: membrane proteins, oxygen-binding proteins: myoglobin and hemoglobin, the protein concentration measurement method I, Bradford methods, the protein concentration measurement method II, Lowry method, BCA method, Biuret method, analytical methods, TCA precipitation, acetone precipitation, immunoprecipitation, concentration of protein solutions, preparative methods, ammonium sulfate precipitation, precipitation with an organic solvent, precipitation with polyethylene glycol, ultrafiltration, dialysis, lyophilization.

Name of Lecturer(s)

Learning Outcomes

1.	Literature review and interpretation of target protein purification
2.	Evaluate, compare and interpret research results
3.	To be able to design and develop protein purification strategies
4.	To be able to relate the general properties of proteins and target protein properties
5.	To be able to understand the use of chromatographic and electrophoretic techniques in separation and protein sequence analysis
6.	To be able to examine and compare the advantages / disadvantages of the methods used

Recommended or Required Reading

1.	Protein Purification Protocols: Paul Cutler
2.	Protein Analysis and Purification: Benchtop Techniques
3.	R. K. Scopes, Protein Purification: Principles and Practice, 3rd ed., Springer- Verlag, (1994) E. L. V. Harris, S. Angal Eds.
4.	Protein Purification Methods; A Practical Approach, 2nd ed., Oxford University Press, (2001) Simpson,R.J

Weekly Detailed Course Contents

Week 1 - Theoretical

Purpose of protein purification and preliminary planning: source and localization of target protein, amount, physicochemical and physical properties

Week 2 - Theoretical

The purpose of protein purification and preliminary planning: the purpose of purification, monitoring of protein purification

Week 3 - Theoretical

Protein saflaştırma stratejisi , Ön Ayırma teknikleri; 1. Hücre parçalama/ Homojenizasyon: Mekanik olmayan Teknikler: ısı ile parçalama, Freze-Thaw, Desikasyon, Osmotik şok, Litik enzimler, Alkali, Deterjanlar vb.

Week 3 - Practice

Cell lysis / Homogenization

Week 4 - Theoretical

Pre-separation techniques (continued) Mechanical methods: Blenders, homagenizers, grinders, Agitation with Abrasives, Liquid and Solid Extrusion, Ultrasonication

Week 4 - Practice

Blenders, homagenizers, grinders, Agitation with Abrasives, Liquid and Solid Extrusion, Ultrasonication

Week 5 - Theoretical

2. Clarification; Centrifugal; Differential Centrifuge, Density Gradient Centrifuge, Membrane Filtration techniques

Week 5 - Practice

2. Clarification; Centrifugal; Differential Centrifuge

Week 6 - Theoretical

3. Concentration of extract: Ultrafiltration, Lyophilization

Week 7 - Theoretical

3. Concentration of the extract: precipitation methods (isoelectric precipitation, precipitation by changing ionic strength, organic solvents, organic polymers and denaturation precipitation)

Week 8 - Intermediate Exam

Protein Isolation and Identification Techniques Midterm Exam

Week 9 - Theoretical

Chromatographic Methods; Introduction to chromatography and basic concepts in liquid chromatography, ion exchange chromatography

Week 10 - Theoretical

Chromatographic Methods; Hydrophobic Interaction Chromatography, Gel Filtration Chromatography

Week 11 - Theoretical

Chromatographic Methods; Affinity chromatography: Biospecific, Immunoaffinity, Lectin affinity, Dye-ligand, Metal-Chelate, Covalent affinity chromatography

Week 12 - Theoretical

Electrophoretic Methods; Principles of electrophoresis, protein electrophoresis; PAGE, SDS-PAGE, IEF, 2D-PAGE. Blotting, Immunustain, Imaging and evaluation

Week 12 - Practice

Protein electrophoresis; PAGE, SDS-PAGE, IEF, 2D-PAGE. Blotting, Immunustain, Imaging and evaluation

Week 13 - Theoretical

Characterization: Functional Characteristic; Techniques to be selected according to biological activity of protein

Week 14 - Theoretical

Purity control, Molecular mass determination and IP determination Amino acid analysis, N-, C-terminal analysis, etc.

Week 15 - Theoretical

Integration and comparison, proteomic technologies and multidimensional techniques

Week 16 - Final Exam

Protein Isolation and Identification Techniques Final Exam

Assessment Methods and Criteria

Type of Assessment	Count	Percent
Midterm Examination	1	%40
Final Examination	1	%60

Workload Calculation

Activities	Count	Preparation	Time	Total Work Load (hours)
Lecture - Theory	14	0	1	21
Lecture - Practice	14	0	1	21
Laboratory	6	3	9	72
Midterm Examination	1	4	1	5
Final Examination	1	5	1	6
TOTAL WORKLOAD (hours)				125

Contribution of Learning Outcomes to Programme Outcomes

	PÇ-1	PÇ-2	PÇ-3	PÇ-4	PÇ-5
OÇ-1	5	5	4	5	5
OÇ-2	5	4	5	4	5
OÇ-3	5	5	5	4	5
OÇ-4	5	4	5	5	4
OÇ-5	5	4	5	5	4
OÇ-6	5	4	5	5	4

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