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Introduction

History

Function

NF1 is a GTPase-activating protein that binds to Ras, a GTPase, to increase the hydrolysis of GTP to GDP. This inactivates the cell signaling of Ras until another GTP can replace the GDP from the cytosol. NF1 and Ras binding is possible in only the conformation of NF1. The mechanism is shown in figure 1 and displays the slow hydrolysis of GTP bound to Ras and the fast hydrolysis of GTP when bound to NF1.

Structure

NF1 is a protein dimer that exists in the conformation and conformation conformation. Each protomer contains a GRD, Sec14-PH, and a GAPex domain located on a HEAT N-C arm. Ras binds to the GRD site with Arg1276 being the critical residue for binding.

Closed conformation

In the conformation, one protomer has its domains shifted by a 130 degree rotation of three separate linkers. That rotation places in an orientation that in the GRD site (Figure 3). Making sterically impossible in the conformation . The conformation can exist naturally without any form of stabilization but will also fall back to the conformation.

Zinc Stabilized

The conformation of NF1 can be stabilized by a zinc ion to prevent the shift back to an conformation. This binding is done between C1032, H1558, and H1576 within the N-HEAT domain, GRD-Sec14-PH linker L2 and is shown in figure #. When zinc stabilizes NF1, it will stay in the conformation and continue to inhibit the binding of RAS.

Open conformation

In the conformation one protomer is shifted due to a 90 rotation. This rotation allows for binding between RAS and the in the GRD site while in the conformation. Allowing for the to occur without any steric hindrance as shown in the conformation.

Conformational Change Linkers

The rotation of the domains between the conformation and conformation of NF1 are conducted by three helical linkers named L1, L2, and L3. The and the undergo rotations to relocate the GRD and Sec14-PH sites. Linker 1 (L1) consists of a loop connected by two helices from L1173-M1215 and is the main contributor in rotation of the GRD domain. The rotation of L1 to the causes N-HEAT ARM alpha helix 48 and GRD helix 49 to extend out, aligning to form a hinge point at G1190. The GRD relocation is assisted by Sec14-PH relocation, which is initiated by Linker 3(L3) from Q1835 to G1852 where the proline rich section of the C-HEAT ARM changes conformation.L1 and L3 move closer to each other in the . Linker 2 (L2) consists of residues G1547-T1565 and begins at helix 63, the final helix of the GRD site, and connects into the short loop of alpha helix 65 of the Sec14-PH domain. The combination of these three linkers are responsible for the conformational shift of the and .

Domains

GRD

The GRD site is represented in cyan in all models. NF1’s main catalytic domain is the GRD active site. Linked structurally to both HEAT ARM’s, it consists of mainly loops and helices. Per protomer, there is one single GRD binding site. In the closed state Ras cannot bind due to a steric hindrance in which Ras clashes with the N-HEAT ARM upon attempting to bind to the GRD site. In its active state GRD can bind Ras. The critical residue within the GRD site is Arg1276.

GAPex-Subdomain

The GAPex subdomain is represented in magenta in all models. The GAPex subdomain of the GRD site lies between the Sec14-PH and GRD catalytic sites. This domain is non-catalytic and structurally consists of various loops and helices. Its main function is to bind SPRED-1, which is a recruiter protein that binds to this subdomain in the cytosol to recruit NF1 to the plasma membrane.

Sec14-PH

The Sec14-PH domain is represented in yellow in all models. The Sec14-PH domain is linked and extends out from the HEAT ARM’s and consists of largely various helices and loops. Its function is a membrane associated domain and holds a largely hydrophobic cavity allowing for binding to the plasma membrane. In the conformation it is blocked by the GRD and is inaccessible to the lipid membrane. In the conformation it becomes exposed and can access the lipid membrane for interaction.

Arginine 1276

Arg1276 is the critical residue within the GRD site needed for proper binding to Ras. The interaction between is only possible in the open confirmation. has making binding of Ras impossible (Figure #).

SPRED 1

SPRED 1 is a protein that binds to the GAPex domain of NF1. Its function recruits the NF1 protein when bound from the cytosol to the plasma membrane. SPRED 1 will bind to the GAPex domain of NF1 in the in the cytosol to recruit NF1 to the plasma membrane. Unlike Ras, SPRED-1 does show the ability to . When bound to the open conformation of NF1 in the cytosol, it may present a different orientation that impacts the recruitment to the plasma membrane. Further research is needed to assess the impact of the function and the changes it may present.

References placeholders

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References

1. ↑ Bergoug M, Doudeau M, Godin F, Mosrin C, Vallee B, Benedetti H. Neurofibromin Structure, Functions and Regulation. Cells. 2020 Oct 27;9(11). pii: cells9112365. doi: 10.3390/cells9112365. PMID:33121128 doi:http://dx.doi.org/10.3390/cells9112365
2. ↑ Bourne HR. G proteins. The arginine finger strikes again. Nature. 1997 Oct 16;389(6652):673-4. doi: 10.1038/39470. PMID:9338774 doi:http://dx.doi.org/10.1038/39470
3. ↑ Kiuru M, Busam KJ. The NF1 gene in tumor syndromes and melanoma. Lab Invest. 2017 Feb;97(2):146-157. doi: 10.1038/labinvest.2016.142. Epub 2017, Jan 9. PMID:28067895 doi:http://dx.doi.org/10.1038/labinvest.2016.142
4. ↑ Lupton CJ, Bayly-Jones C, D'Andrea L, Huang C, Schittenhelm RB, Venugopal H, Whisstock JC, Halls ML, Ellisdon AM. The cryo-EM structure of the human neurofibromin dimer reveals the molecular basis for neurofibromatosis type 1. Nat Struct Mol Biol. 2021 Dec;28(12):982-988. doi: 10.1038/s41594-021-00687-2., Epub 2021 Dec 9. PMID:34887559 doi:http://dx.doi.org/10.1038/s41594-021-00687-2
5. ↑ Naschberger A, Baradaran R, Rupp B, Carroni M. The structure of neurofibromin isoform 2 reveals different functional states. Nature. 2021 Nov;599(7884):315-319. doi: 10.1038/s41586-021-04024-x. Epub 2021, Oct 27. PMID:34707296 doi:http://dx.doi.org/10.1038/s41586-021-04024-x
6. ↑ Sabatini C, Milani D, Menni F, Tadini G, Esposito S. Treatment of neurofibromatosis type 1. Curr Treat Options Neurol. 2015 Jun;17(6):355. doi: 10.1007/s11940-015-0355-4. PMID:25917340 doi:http://dx.doi.org/10.1007/s11940-015-0355-4