Keratins: Difference between revisions

<Structure load='3tnu' size='300' frame='true' align='right' caption='Fig. 1. Crystal structure of the 2B helical domain of coiled-coil dimer of type I keratin K14 (chain A) and type II keratin K5 (chain B) (residues Ser332-Gly421 of K14 and Thr382-Gly476 of K5). PDB ID: 3TNU.' scene='Insert optional scene name here' />

Keratin is the name given to a large family of homologous proteins that have a filamentous (fibrous) structure. These proteins are expressed in epithelial cells and in epidermal cells where they they are assembled forming cytoskeletal structures within the cell and epidermal derivatives such as hair, nail and horn <ref>PMID:18461349</ref>.  

The keratins represent the largest branch within the super-family of intermediate-filament (IF) proteins <ref name="PMID18083519">PMID:18083519</ref> <ref>PMID:19587451</ref>. Keratins are grouped into two families termed as type I and type II keratins based on their sequence homology <ref name="PMID6191871">PMID:6191871</ref>. Similarly, other IF proteins are also grouped into families termed consecutively as types III, IV, V and VI IF proteins, based on their sequence homology <ref>PMID:8982454</ref>. These families include desmin, vimentin, neurofilament protein and GFAP that are expressed in specific tissues and cell types <ref name="PMID18083519" />. The IF family of lamins are located on the nuclear lamina and are ubiquitously expressed <ref name="PMID18083519" />.  

* Microfilaments composed of actin subunits  

* Microtubules composed of tubulin subunits

The basic building block of each intermediate filament is a dimer of a coiled-coil pair of IF proteins. Each keratin filament is assembled as a hetero-dimer of a type I keratin coiled together with a type II keratin. <ref name="PMID6191871">PMID:6191871</ref>. Other types of IFs are mostly composed of homo-dimers <ref name="PMID18083519" />.  

In humans there are 54 functional genes that code for keratins <ref>PMID:16831889</ref> <ref name="PMID15085952">PMID:15085952</ref>. The first sequences of human keratin cDNAs revealed that there are two distinct but homologous keratin families <ref name="PMID6191871" /> <ref name="PMID6186381">PMID:6186381</ref>. These two distinct types were named as Type I keratin and Type II keratin <ref name="PMID6191871" />.  

Human genome sequencing revealed that type I and type II keratin genes are located in two clusters each of which includes 27 genes on chromosome 17q21 and on chromosome 12q13 respectively <ref name="PMID15085952" /> <ref>PMID:17428470</ref>. The juxtaposed location of the genes indicate that these gene clusters evolved by a series of gene duplication events.

Determination of the sequences of type I and type keratins revealed that the two types of keratins have a central ~310 residue long segment that share ~30% homology, but the amino and carboxy terminal regions of these proteins show great diversity <ref name="PMID6186381" />. Consistent with the initial observations, sequencing of keratins and other intermediate filament proteins showed that all IF proteins have a conserved central domain and widely divergent amino and carboxy terminal regions <ref>PMID: 17521629</ref>.  

Sequencing and two dimensional gel electrophoresis of the complete family of keratins revealed that the type I and type II keratins differ in their size and isoelectric points <ref name="PMID19422428">PMID:19422428</ref> <ref>PMID:18461349</ref>. Type I keratins are generally smaller (average length 460 aa's), and acidic (isoelectric point 4.4-5.4), while type II keratins are longer (average length 545 aa's) and basic (isoelectric point 5-8.3). As noted, the size differences among keratins result from differences in the amino and carboxy terminals of the proteins <ref name="PMID6191871" />.

[[Image:Keratin-secondary-structure-1000px.png|600px|right|thumb| Fig. 2. The locations of the &alpha;-helical domains (1A, 1B, 2A and 2B) in the central rod of a keratin subunit.]]

Analysis of the first cytoskeletal keratin sequence revealed that this protein contains a central domain of ~310 residues that was predicted to be mostly in &alpha;-helix conformation <ref name="PMID6186381" />. By comparative analysis of the predicted structures of a type I keratin, a type II keratin, desmin and vimentin, Hanukoglu and Fuchs suggested that all IF proteins have a central ~310 residue domain that contains four segments in &alpha;-helical conformation that are separated by three short linker segments predicted to be in beta-turn conformation <ref name="PMID6191871" />. This model has been confirmed by analysis of the crystal structure of segments of keratin coiled-coil <ref name="PMID22705788" >PMID:22705788</ref>.  

The structures of the head and tail domains of keratins are highly variable and have not been elucidated. Based on their sequences, these domains are predicted to be non-helical, probably forming globular structures that participate in interactions between subunits and other proteins in the scaffold of cellular cytoskeleton <ref name="PMID19422428" />.

Keratin fibers are difficult to solubilize and so far it has not been possible to crystallize a whole keratin or a combination of keratin polymers. In the face of this difficulty, soluble segments of keratins have been generated both by proteolytic digestion and gene engineering to study the structural properties of keratins <ref name="PMID17521629">PMID:17521629</ref> .  

As noted above, keratin filaments are composed of hetero-dimers. To express the long 2B segment of hetero-dimer of keratins K5 and K14, Lee et al. transformed two cDNAs into E. coli, isolated the heteromeric complex, and crystallized it. Structural analysis revealed a coiled-coil hetero-dimer structure of K5 and K14 intertwined around one another. These findings establish that keratin filament is composed of a coiled-coil hetero-dimer wherein the 2B segments are intertwined in parallel <ref name="PMID19422428" />.  

All evidence to date indicates that the basic unit of a keratin filament is a left-handed hetero-dimer of a matched pair of keratins aligned in parallel. The ~10 nm wide keratin filament is assembled in several steps <ref name="PMID19422428" />:

<Structure load='3TNU' size='600' frame='true' align='right' caption='Fig. 3. Coiled-coil dimer of type I keratin K14 and type II keratin K5.' />

The crystal structure of coiled-coil 2B helical domains of keratins K5 and K14, have revealed the bonds that are involved in tight binding of the two subunits <ref name="PMID19422428" />.

In the 2B domains of keratins type I K14 and type II K5 shown in Fig. 3, there are two and a single cysteine respectively. These cysteines are far apart and cannot form disulfide bridges.  

* <scene name='55/559109/Cysteines/1'>Click here to see the locations of the Cys in Fig. 3.</scene> (Wait a few moments for change of scene)

* <scene name='55/559109/Acidic-residues/1'>Click here to see the acidic residues, Asp and Glu in Fig. 3.</scene>

Both acidic and basic residues are seen to protrude mostly towards the outside surface of the two keratins and hardly in the space between the two keratins. The contact surface between the two keratins in a coiled-coil is located between the two keratins. Thus, the charged residues do not play a predominant role in the formation of the coiled-coil. In the K14-K5 dimer only 3-4 residues are involved in inter-strand interactions. Nonetheless, these residues are essential for normal function of keratin <ref name="PMID19422428" />.

* <scene name='55/559109/Hydrophobic-aas/1'>Click here to see the hydrophobic residues in Fig. 3.</scene>

As the two chains of keratins are intertwined in parallel, the contact points along the entire coiled-coil represents a seam along the two proteins. Coiled-coil structures are found in many types of proteins. In two-chained coiled-coil proteins hydrophobic residues appear in a periodic pattern that has been named a heptad-repeat <ref name="PMID15837514">PMID:15837514</ref> . In a regular &alpha;-helix there are 3.6 residues per turn of the helix. In a left-handed coiled-coil there are 3.5 residues per turn. Thus, in a two chained coiled-coil there is a repeat pattern of seven residues that are represented by the letters a-b-c-d-e-f-g. Residues a and d in this pattern  are hydrophobic. These two residues define a hydrophobic flank for each protein. This periodic pattern was first reported on both type I and type II wool keratins <ref name="PMID697726">PMID:697726</ref> and later observed on cytoskeletal keratins as well <ref name="PMID6191871" />. The crystal structures of the 2B segment of keratins K14 and K5 provided final confirmation for the role of these hydophobic residues in coiled-coil formation <ref name="PMID19422428" />.