ABCB10 p.Leu46Gln
| Predicted by SNAP2: | A: N (53%), C: N (66%), D: D (80%), E: D (75%), F: D (53%), G: D (75%), H: D (71%), I: N (78%), K: D (75%), M: N (78%), N: D (71%), P: D (75%), Q: D (63%), R: D (75%), S: D (63%), T: D (59%), V: N (78%), W: D (66%), Y: D (63%), |
| Predicted by PROVEAN: | A: N, C: N, D: N, E: N, F: N, G: N, H: N, I: N, K: N, M: N, N: N, P: N, Q: N, R: N, S: N, T: N, V: N, W: N, Y: N, |
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[hide] Targeting, import, and dimerization of a mammalian... J Biol Chem. 2004 Oct 8;279(41):42954-63. Epub 2004 Jun 23. Graf SA, Haigh SE, Corson ED, Shirihai OS
Targeting, import, and dimerization of a mammalian mitochondrial ATP binding cassette (ABC) transporter, ABCB10 (ABC-me).
J Biol Chem. 2004 Oct 8;279(41):42954-63. Epub 2004 Jun 23., [PMID:15215243]
Abstract [show]
ATP binding cassette (ABC) transporters are a diverse superfamily of energy-dependent membrane translocases. Although responsible for the majority of transmembrane transport in bacteria, they are relatively uncommon in eukaryotic mitochondria. Organellar trafficking and import, in addition to quaternary structure assembly, of mitochondrial ABC transporters is poorly understood and may offer explanations for the paucity of their diversity. Here we examine these processes in ABCB10 (ABC-me), a mitochondrial inner membrane erythroid transporter involved in heme biosynthesis. We report that ABCB10 possesses an unusually long 105-amino acid mitochondrial targeting presequence (mTP). The central subdomain of the mTP (amino acids (aa) 36-70) is sufficient for mitochondrial import of enhanced green fluorescent protein. The N-terminal subdomain (aa 1-35) of the mTP, although not necessary for the trafficking of ABCB10 to mitochondria, participates in the proper import of the molecule into the inner membrane. We performed a series of amino acid mutations aimed at changing specific properties of the mTP. The mTP requires neither arginine residues nor predictable alpha-helices for efficient mitochondrial targeting. Disruption of its hydrophobic character by the mutation L46Q/I47Q, however, greatly diminishes its efficacy. This mutation can be rescued by cryptic downstream (aa 106-715) mitochondrial targeting signals, highlighting the redundancy of this protein's targeting qualities. Mass spectrometry analysis of chemically cross-linked, immunoprecipitated ABCB10 indicates that ABCB10 embedded in the mitochondrial inner membrane homodimerizes and homo-oligomerizes. A deletion mutant of ABCB10 that lacks its mTP efficiently targets to the endoplasmic reticulum. Quaternary structure assembly of ABCB10 in the ER appears to be similar to that in the mitochondria.
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No. Sentence Comment
9 Disruption of its hydrophobic character by the mutation L46Q/ I47Q, however, greatly diminishes its efficacy.
X
ABCB10 p.Leu46Gln 15215243:9:56
status: NEW229 We removed one hydrophobic residue pair with the double mutation L46Q/I47Q.
X
ABCB10 p.Leu46Gln 15215243:229:65
status: NEW243 Mutation MMCC n Wild type 0.77 Ϯ 0.02 12 aa 36-70-R41A 0.71 Ϯ 0.04 12 aa 36-70-R55A 0.76 Ϯ 0.03 12 aa 36-70-R41A-R55A 0.74 Ϯ 0.03 28 aa 36-70-R41P 0.73 Ϯ 0.02 19 aa 36-70-A42P 0.75 Ϯ 0.03 16 ing-The flanking thirds of the mTP, aa 1-35 and 71-105, only partially rescue the effect of the L46Q/I47Q mutations on aa 36-70, demonstrating that the Leu46 -Ile47 hydrophobic pair of the mTP of ABCB10 is essential for its mitochondrial targeting (Fig. 4C).
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ABCB10 p.Leu46Gln 15215243:243:324
status: NEW246 The R41A/L46Q/I47Q mutations were carried out in full-length ABCB10 (715 aa) fused to EGFP (Fig. 4E).
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ABCB10 p.Leu46Gln 15215243:246:9
status: NEW257 B, the L46Q/I47Q mutant is additionally susceptible to reduction of targeting efficiency by R41A mutation.
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ABCB10 p.Leu46Gln 15215243:257:7
status: NEW258 The asterisk indicates significant difference from aa 36-70-L46Q/I47Q.
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ABCB10 p.Leu46Gln 15215243:258:60
status: NEW259 C and D, rejoining aa 1-35 and 71-105 to aa 36-70-L46Q/I47Q and aa 36-70-L46Q/I47Q/R41A partially restores mitochondrial targeting.
X
ABCB10 p.Leu46Gln 15215243:259:50
status: NEWX
ABCB10 p.Leu46Gln 15215243:259:73
status: NEW262 E, fusing aa 106-715 to aa 1-105-L46Q/I47Q/R41A restores mitochondrial targeting to that of wild-type ABCB10.
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ABCB10 p.Leu46Gln 15215243:262:33
status: NEW316 On the contrary, the hydrophobic residues responsible for Tom20 interaction must meet stringent accessibility and location requirements, since all nine additional hydrophobic pentapeptide motifs of the aforementioned type introduced by the flanking regions failed to completely rescue the L46Q/I47Q mutation.
X
ABCB10 p.Leu46Gln 15215243:316:289
status: NEW