BNG 331 Cell-Tissue Material Interactions. Proteins

Transcription

1 BNG 331 Cell-Tissue Material Interactions Proteins Monday, April 8, 2013

2 Course update A couple notes on reading papers Review LBL guidelines Grading: only individual differences will be in speaking style (no peer evaluation!) Journal article discussion Wednesday will simulate an LBL! For the paper, list any questions that came to mind as you read it Please think about the discussion questions (distributed in class Friday)

3 Copolymers (from last class) Use of copolymers to mix material properties polycaprolactone poly(ethylene glycol) (PCL-PEG): PEG cheap hydrophilic good mechanics PCL biodegradable the ester is water-degradable

4 LBL groups and dates (on website) LBL 1 (F 4/19): Brian Choi, Andrew Thistle, John Bocchino, Phil Kemp, Tim Hersey LBL 2 (F 4/26): Catherine Tjan, Kyra Burnett, Joe Maher, Peter Aspinall LBL 3 (F 5/3): Adam Dear, Alex Kazarian, Sean Day, Taylor Tebano, Ben Stephens <Friday, 5/10 Steinmetz Symposium, no classes> LBL 4 (F 5/17): John Paul Dumas, Tanner Jones, Meagan Carnes, Amy Loya LBL 5 (F 5/24): Kristin O Connor, Brian Wilson, Chris Sullivan, Stanley Gelin LBL 6 (F 5/31): Sarah Bradner, Kyle Chau, Kim Derosier, Andrew Gioe, Michael Grabarits

5 Proteins are biopolymers Recall this slide from the previous lecture: proteins (polypeptide, amino acid) nucleic acids (polypeptide, amino acid) carbohydrates (polysaccharide, monosaccharide) lipids (not true polymers, though are sometimes referred to as such)

6 Biological functions of proteins While nowhere near exhaustive, the role of proteins in the body include: As the primary structural component of extracellular matrix (ECM) for many/most tissues Catalysis of thousands of important chemical reactions essential to life Regulation of gene transcription Conversion of food to energy Changes in the levels or function (via mutations) of proteins account for many disease processes Osteogenesis imperfecta type 1 collagen

7 Structure of proteins let s take a look at these one at a time

8 Primary (1 ) structure The amino acid is the monomer or building block for proteins Based on the side chain (R), can divide AAs into 4 groups: Nonpolar (hydrophobic) Polar Positively charged Negatively charged However, the above groups are not mutually exclusive (let s see what we mean)

9 20 standard amino acids structures Non-aromatic aromatic containing rings of conjugated carbons

10 20 standard amino acids properties

11 Primary (1 ) structure peptides Two amino acids are covalently bound via an enzymecatalyzed condensation reaction: This molecule is now a peptide

12 Primary (1 ) structure peptides Oligopeptides versus polypeptides: amino acid (n = 1) oligopeptide (n < 20) polypeptide (n > 30-50) However, generally n >> 50 for proteins let s do a quick exercise to see how many amino acids are in insulin (5808 g/mol)? To do this, let s introduce the dalton (Da)

13 Insulin 1 structure n = = 51 amino acids

14 Albumin 1 structure M = 66,463 Da; n = 583 amino acids

15 Secondary (2 ) structure Secondary (non-covalent) interactions between AAs (prmarily H bonding, hydrophobic) cause the 1 structure to fold, bend, and coil to give it a 3D structure called a conformation For example, coiling and bending caused by H bonding between a carbonyl group (-C=O) of one peptide and a secondary amine (- NH) of another The bending is constrained by the rotational angle of the covalent bonds α-helices and β-sheets are the most thermodynamically stable, and thus, the most common α-helix: an H bond is formed between n th and (n + 4) th AAs β-sheet: H bonds between at least two adjacent strands

16 Tertiary (3 ) structure The overall 3D structure of one polypeptide chain, i.e., of one subunit 3 structure is governed by interactions between distant sections of the polypeptide chain Not only H bonding of carbonyl/amino groups, but other types of 2 (non-covalent) bonding (ionic bonds, hydrophobic interactions) between the variable side groups of AAs Goal of the subunit: maximize polar group exposure and minimize hydrophobic group exposure to the aqueous environment Example 1 Example 1 (hydrophobic interactions)

17 Quaternary (4 ) structure heme groups with bound Fe atoms for oxygenation Interactions between separate subunits (polypeptide chains) in multi-chain proteins Same types of bonds stabilize the structure (now between chains rather tan within a chain) Hemoglobin (each chain is a different subunit)

18 Common proteins found in vivo

19 Collagen overview The most abundant protein in the human body (~25% by mass) 19 distinct types have been identified; some common examples: Type I (~300 kda): most abundant, found in bone, tendons, ligaments, skin, and many organs strong in tension Type II (~400 kda): primary structural component of articular and hyaline cartilage Type X (~350 kda): present in hypertrophic and mineralizing cartilage All collagens are made up three coiled alpha polypeptides let s take a closer look at the structure

20 Collagen structure another view: arr/collagen_structure.html

21 Elastin overview A major component of tissues that require elasticity to function can you name a few? Arteries: particularly common in large, highly elastic blood vessens (such as the aorta) Elastic ligaments Skin bladder Rich in glycine and proline Hydrophobic AAs make up about 50% of the primary structure

22 Elastin structure The individual unit of elastin and product of the ELN gene is tropoelastin (~72 kda) After secretion, individual tropoelastin chains are covalently crosslinked to form highly insoluble networks of elastic fibers individual tropoelastin Baldock et al. PNAS 2011 tropoelastin crosslinked elastin sigmaaldrich.com

23 Fibronectin Extracellular matrix (ECM) noncollagenous adhesive proteins (~440 kda) which organize matrix and allow/enable cells to attach to it RGD, derived from fibronectin, is a very commonly used amino acid sequence for cell adhesion Very important for endothelial cell function (we will see this Monday) Fibronectin-mutant mice die as embroys due to the inability to form blood vessels wikipedia.com

24 Fibronectin and fibrinogen Fibronectin has sticky arms that can bind heparin, collagen, cells (via integrins) and even more fibronectin Fibrinogen (~340 kda) is a much more complex structure A plasma protein is present in blood at up to 3 mg/ml Serves as the final target of the coagulation cascade

25 Overall: Properties and functions of proteins are directly related to the intramolecular interactions resulting from primary structures

26 Importance of proteins for biomaterials? 3 major ways (amongst others!) in which proteins are revelant in the field of biomaterials: 1) Surface modification of biomaterials with proteins or peptides: Typically uses adsorption (proteins) or covalent coupling (peptides) to the surface cell adhesion Engler AJ, et al. Cell, using proteins or fragments (could also use fibronectin) using peptides (non-rgd sequences are also used) 2) Loading of proteins (e.g., growth factors, chemotherapeutics) into biomaterials (e.g., biodegradable microspheres) for controlled release A challenge is maintenance of the bioactivity/stability of encapsulated proteins 3) Interactions between biomaterial surfaces and in vivo proteins following implantation into the body The focus of Friday s lecture (Professor Currey)

27 Questions?