This web page was produced as an assignment for Genetics 564, an undergraduate course at UW-Madison.
What is a protein?
Proteins are what make up who we are, with the largest visible protein being keratin (our hair). How many proteins are there, where do they come from, and how are they made? It is estimated that humans have from 1,200,000- 10,000,000 different proteins. These protein come from the genes that make up our DNA. A common phenomenon that explains how DNA goes to protein is termed the central dogma (Figure1). Protein formation is focused on translation where mRNA is translated by ribosomes. Here tRNAs work with ribosomes coding for the specific amino acids needed to make the specific protein from the mRNA. After completion of translation various other proteins aid in folding the amino acid strand into the correct protein. We'll further explore the FLNB protein coded from the FLNB gene that aids in proper development.
The FLNB Protein :
The FLNB protein, commonly called filamin B, is transcribed and translated from the FLNB gene located on chromosome 3p14.3. The longest FLNB protein is composed of 2633 amino acids and is the isoform that I used throughout this website. Other known isoforms of the FLNB protein are discussed below. FLNB in an important protein for proper development before birth and variations in FLNB leads to severe defects and/ or lethality (2). FLNB function within the cell is to connect the actin cytoskeleton filaments to the plasma membrane which allows cells to change shape and move (1-2). The actin filaments are arranged in a lattes pattern all over the cell to create the cytoskeleton (Figure 2). The actin binding domain at the N-termini of the FLNB protein binds to the actin filaments and the C-terminus interacts with plasma membrane proteins allowing actin to be bound to the plasma membrane by FLNB.
FLNB has great importance in proper bone development and joint stability, though the science behind why it does is still unclear. What we do know is that in the case of Larsen Syndrome and other genetic disorders, such as spondlyocarpotarsal syndrome and atelosteogenesis I and III, located on the FLNB gene/ protein cause bone and joint defects (3). In Larsen Syndrome the affected FLNB protein, by a missense variant, causes an abnormal protein to form (2). This abnormal protein causes the actin cytoskeleton to be disorganized within the cell. |
What are the known variants in FLNB that cause Larsen Syndrome?
The resulting cause of Larsen Syndrome is when one or several missense variants along the FLNB protein occurs. A missense variant is when a base pair change occurs in the DNA, during transcription, which codes for a different amino acid that alters the composition of the protein. The most common missense variant in Larsen Syndrome patients is where a G (guanine) is changed to an A (3). Two small scale studies have been performed that analyzed the DNA of patients with Larsen Syndrome to try to determine the most common variants related with the disorder or where along the FLNB protien they most likely occur. One study looked at 8 patients and identified 5 causative variants (4) (Figure 3: Blue arrows) and the other looked at 20 patients and identified 8 new variants (3) (Figure 3: Gold arrows). These two studies have shown that variants resulting in Larsen Syndrome appear to be along the second CH domain and from filamin 13-17 domains (3). With larger studies they could identify many more mutations along the FLNB protein that result in Larsen Syndrome as with each study performed more and more new variants are being discovered.
What is an Isoform?
An isoform is when there are different forms of a protein produced for the same or different genes (5). This is done through alternative splicing of the mRNA, where different exons can be expressed, during transcription of the central dogma (Figure 1). The isoforms of the FLNB protein are all from the same FLNB gene located on chromosome 3p14.3.
How many FLNB isoforms are there?
There is some variation when it comes to how many isoforms there are of the FLNB protein. Uniprot lists 9 FLNB isoforms where as NCBI identifies 4. I decided to look at the four isoforms discovered on NCBI's database because their genetic sequences were identified via FASTA (found below), where as not all of the isoforms listed on Uniprot were. I was then able to individually input each FLNB isoform's amino acid sequence into the SMART database to determine each isoforms domains and differences (Figure 4). On SMART be sure the have the "genomic" smart mode chosen before submitting the sequence. The isoform used throughout this website was #1for it contained the largest amount of amino acids, 2633, therefore no information would be missed when further analyzing the FLNB protein.
FLNB Isoform References:
Few quick facts about the FLNB protein:
Using ExPASy, the molecular weight and pI of the FLNB protein (2633 amino acids) are:
- MW: 281634.79 Da
- pI: 5.46
This was achieved by entering in FLNB protein FASTA sequence (bottom of Protein Homology) into the ExPASy Compute pI/Mw tool. pI stands of isoelectric point. The isoelectric point is the pH at which the FLNB protein surface or molecule carries no net electrical charge (7).
- MW: 281634.79 Da
- pI: 5.46
This was achieved by entering in FLNB protein FASTA sequence (bottom of Protein Homology) into the ExPASy Compute pI/Mw tool. pI stands of isoelectric point. The isoelectric point is the pH at which the FLNB protein surface or molecule carries no net electrical charge (7).
References:
1.)"Uniprot:O75369 (FLNB_HUMAN)". Web. May 10, 2014. http://www.uniprot.org/uniprot/O75369
2.) "Genetics Home Reference: Larsen Syndrome". Web. January 26, 2014. http://ghr.nlm.nih.gov/condition/larsen-syndrome
3.) Bicknell, L. S., et. al., (2007). "A molecular and clinical study of Larsen Syndrome caused by mutations in FLNB". Journal of Medical Genetics, 44(1), doi:10.1136/jmg.2006.043687
4.) Krakow, D., et. al., (2004). "Mutations in the gene encoding filamin B disrupt vertebral segmentation, joint formation and skeletogenesis". Nature Genetics, 36(4), doi:10.1038/ng1319
5.) "Genetics Home Reference: Isoforms". Web. May 11, 2014. http://ghr.nlm.nih.gov/glossary=isoforms
6.) "ExPASy: FLNB_HUMAN (075369)". Web. May 9, 2014. http://web.expasy.org/cgi-bin/compute_pi/pi_tool1?O75369@1-2602@average
7.) "Isoelectric Point". Web. May 14, 2014. http://www.princeton.edu/~achaney/tmve/wiki100k/docs/Isoelectric_point.html
2.) "Genetics Home Reference: Larsen Syndrome". Web. January 26, 2014. http://ghr.nlm.nih.gov/condition/larsen-syndrome
3.) Bicknell, L. S., et. al., (2007). "A molecular and clinical study of Larsen Syndrome caused by mutations in FLNB". Journal of Medical Genetics, 44(1), doi:10.1136/jmg.2006.043687
4.) Krakow, D., et. al., (2004). "Mutations in the gene encoding filamin B disrupt vertebral segmentation, joint formation and skeletogenesis". Nature Genetics, 36(4), doi:10.1038/ng1319
5.) "Genetics Home Reference: Isoforms". Web. May 11, 2014. http://ghr.nlm.nih.gov/glossary=isoforms
6.) "ExPASy: FLNB_HUMAN (075369)". Web. May 9, 2014. http://web.expasy.org/cgi-bin/compute_pi/pi_tool1?O75369@1-2602@average
7.) "Isoelectric Point". Web. May 14, 2014. http://www.princeton.edu/~achaney/tmve/wiki100k/docs/Isoelectric_point.html