title: Sequence Motifs in Transit Peptides Act as Independent Functional Units and Can Be Transferred to New Sequence Contexts.
abstract: A large number of nuclear-encoded proteins are imported into chloroplasts after they are translated in the cytosol. Import is mediated by transit peptides (TPs) at the N termini of these proteins. TPs contain many small motifs, each of which is critical for a specific step in the process of chloroplast protein import; however, it remains unknown how these motifs are organized to give rise to TPs with diverse sequences. In this study, we generated various hybrid TPs by swapping domains between Rubisco small subunit (RbcS) and chlorophyll a/b-binding protein, which have highly divergent sequences, and examined the abilities of the resultant TPs to deliver proteins into chloroplasts. Subsequently, we compared the functionality of sequence motifs in the hybrid TPs with those of wild-type TPs. The sequence motifs in the hybrid TPs exhibited three different modes of functionality, depending on their domain composition, as follows: active in both wild-type and hybrid TPs, active in wild-type TPs but inactive in hybrid TPs, and inactive in wild-type TPs but active in hybrid TPs. Moreover, synthetic TPs, in which only three critical motifs from RbcS or chlorophyll a/b-binding protein TPs were incorporated into an unrelated sequence, were able to deliver clients to chloroplasts with a comparable efficiency to RbcS TP. Based on these results, we propose that diverse sequence motifs in TPs are independent functional units that interact with specific translocon components at various steps during protein import and can be transferred to new sequence contexts.
authors.affiliation: Division of Integrative Biosciences and Biotechnology (D.W.L., S.W., I.H.) and Department of Life Sciences (K.R.G., I.H.), Pohang University of Science and Technology, Pohang 790-784, Korea.
authors.forename: Dong Wook
authors.initials: DW
authors.lastname: Lee
authors.affiliation: Division of Integrative Biosciences and Biotechnology (D.W.L., S.W., I.H.) and Department of Life Sciences (K.R.G., I.H.), Pohang University of Science and Technology, Pohang 790-784, Korea.
authors.forename: Seungjin
authors.initials: S
authors.lastname: Woo
authors.affiliation: Division of Integrative Biosciences and Biotechnology (D.W.L., S.W., I.H.) and Department of Life Sciences (K.R.G., I.H.), Pohang University of Science and Technology, Pohang 790-784, Korea.
authors.forename: Kyoung Rok
authors.initials: KR
authors.lastname: Geem
authors.affiliation: Division of Integrative Biosciences and Biotechnology (D.W.L., S.W., I.H.) and Department of Life Sciences (K.R.G., I.H.), Pohang University of Science and Technology, Pohang 790-784, Korea ihhwang@postech.ac.kr.
authors.forename: Inhwan
authors.initials: I
authors.lastname: Hwang
mesh_terms: D020816:Amino Acid Motifs; D000595:Amino Acid Sequence; D061146:Chlorophyll Binding Proteins / Q000737:chemistry; D057927:Hydrophobic and Hydrophilic Interactions; D008969:Molecular Sequence Data; D010455:Peptides / Q000737:chemistry*; D021381:Protein Transport; D011523:Protoplasts / Q000378:metabolism; D012273:Ribulose-Bisphosphate Carboxylase / Q000737:chemistry
chemical_list: D061146:Chlorophyll Binding Proteins; D010455:Peptides; D012273:Ribulose-Bisphosphate Carboxylase