SNAT3: structure and pathogenic variants

Sodium-coupled neutral amino acid transporter 3 (SNAT3) is encoded by SLC38A3 and it primarily transports glutamine across the cell membrane in a sodium coupled manner.

Model

Shown is a homology model of SNAT3 based upon the structure of Zebrafish SNAT9 (SLC38A9 encoded) (see methods below for more). This is a cytosol open model. TM1 and TM6 which have a kink would straighten opening up the extracellular side.

Putative organophosphate binding pockets

There are some patches with strong positive charge on cystoplasmic face, a not uncommon feature on the cytosolic leaf of the plasma membrane, and are involved in phospholipid or PIP3 binding —the latter to allow localisation in certain regions such as at synapses. However, without empirical data, the identity of the bound organophosphate cannot be safely ascertained. Consequently, this model has simply three phosphates, which have been docked giving a strong binding energy.

Mutations:

• R208G (wild type/mutant/overlay) —residue in a highly cationic cytosolic pocket, which very likely binds an organophosphate. The mutation results in a +3 kcal/mol increase in binding energy to phosphate.
• splice variant (L374∗) (remnant/missing) — truncation results in a non-functional and likely unfolded protein
• A296T (wild type/mutant/overlay) — in hinge region of TM6, any mutation there is undoubtably going to affect the closing of the cytosolic opening
• S350∗ (remnant/missing) — truncation results in a non-functional and likely unfolded protein
• L374∗ (remnant/missing) — truncation results in a non-functional and likely unfolded protein
• T375P (wild type/mutant/overlay) — prolines break α-helices, the structure of this TM helix is highly compromised (+20 cal/mol). This is stronger than all the destabilising mutation present in gnomAD (highest: R189G at +16 kcal/mol).
• P387Q (wild type/mutant/overlay) —this is slightly destabilising (+3 kcal/mol) and part of the cationic cytosolic pocket of R208, but majorly it is a on a loop on the cytoplasmic opening and the change to a more rigid and bulkier residue may affect the closing of the cytosolic opening.
• W404∗ (remnant/missing) — truncation results in a non-functional and likely unfolded protein

gnomAD

Analysis of gnomAD mutations shows that this protein is highly intolerant of destabilising mutations.

In gnomAD only A327T is homozygous, but it is a conservative change on a structural TM helix, resulting in only +1.9 kcal/mol destabilisation (neutral) —likewise the >1e-4 frequency variants I232T, A327T, V412I, T458M and L481M are neutral. Many mutation occur in regions absent in this model, for example, A3T, T458M, N503S, P24L, R40W, but the low homology would suggest that these regions do not play a critical role. Missense mutations are uncommon in this protein and the most frequent variant that is destabilising (+8 kcal/mol) is E179V at an allele frequency of 5e-5. Only 5 variants are have a ∆∆G between +5 and +10 and only 2 over +10.

Further info

More information, models and code used can be found at github.com/matteoferla/SLC38A3_analysis.