Preparation, Crystallization, and X-ray Data Collection of Archaeal Oligopeptide Permease A

Yokoyama, Hideshi; Kamei, Nanami; Konishi, Koji; Hara, Kodai; Ishikawa, Yoshinobu; Matsui, Ichiro; Forterre, Patrick; Hashimoto, Hiroshi

doi:10.1134/s1063774521070221

Cited by 2 publications

(6 citation statements)

References 35 publications

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“…Among 815 amino acid residues, residues 30–753 of TkOppA that lack an N‐terminal signal peptide and C‐terminal threonine‐rich and membrane‐spanning regions were used. TkOppA (residues 30–753) fused with a C‐terminal 6 × His tag was expressed as a soluble protein in E. coli and purified as described previously 28 . Analyses of ESI–TOF–MS and N‐terminal amino acid sequencing indicated that the purified His‐tagged OppA consisted of residues 30–753 and VEHHHHHH, and an initial methionine was mainly processed 28 .…”

Section: Resultsmentioning

confidence: 99%

“…TkOppA (residues 30–753) fused with a C‐terminal 6 × His tag was expressed as a soluble protein in E. coli and purified as described previously 28 . Analyses of ESI–TOF–MS and N‐terminal amino acid sequencing indicated that the purified His‐tagged OppA consisted of residues 30–753 and VEHHHHHH, and an initial methionine was mainly processed 28 . In order to remove endogenous peptides, which were often observed in the binding pocket of OppA, 6,23 the protein solution was partially unfolded in guanidinium‐HCl (GuHCl) and refolded using Ni‐NTA agarose resin to generate ligand‐free OppA 36 .…”

Section: Resultsmentioning

confidence: 99%

“…The induced cells were centrifuged and stored at −80°C. Using the cells, the SeMet derivative TkOppA_4mut was purified with the same protocol as described previously 28 …”

Section: Methodsmentioning

confidence: 99%

“…Wild-type TkOppA (residues 30-753) was expressed and purified as described previously. 28 To prepare the expression vector pET-26b (Novagen) encoding a I302M/L446M/L557M/L636M quadruple mutant of TkOppA (residues 30-753), which is hereafter referred to as TkOppA_4mut, the following four forward or reverse primers including mutation sites (bolded) were used: I302M_rev, 5 0 -ATTGTGCTCGTCCATGAATTTGGCTATT-3 0 ; L446M_fwd, 5 0 -GCCA TCCAGTACATGGTTAACAGGGCTT-3 0 ; L557M_rev, 5 0 -GCCGAG CTTCTTCATGACTTCCTCAACT-3 0 ; L636M_fwd, and 5 0 -CAGAGGG TTACAATGGAGGAGGTTCTCA-3 0 . Polymerase chain reaction was carried out using pET-26b encoding TkOppA (residues 30-753) as a template and KOD plus polymerase (Toyobo).…”

Section: Preparation Of Tkoppamentioning

confidence: 99%

“…28 Analyses of ESI-TOF-MS and N-terminal amino acid sequencing indicated that the purified His-tagged OppA consisted of residues 30-753 and VEHHHHHH, and an initial methionine was mainly processed. 28 In order to remove endogenous peptides, which were often observed in the binding pocket of OppA, 6,23 the protein solution was partially unfolded in guanidinium-HCl (GuHCl) and refolded using Ni-NTA agarose resin to generate ligand-free OppA. 36 Then, the solution was mixed with the peptide bradykinin (RPPGFSPFR) and crystallized in order to elucidate interactions with the specific peptide rather than endogenous peptides.…”

Section: Structure Determination Of Tkoppamentioning

confidence: 99%

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Structural basis for peptide recognition by archaeal oligopeptide permease A

et al. 2022

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Oligopeptide permease A (OppA) plays an important role in the nutrition of cells and various signaling processes. In archaea, OppA is a major protein present in membrane vesicles of Thermococcales. Because there being no crystal structures of archaeal OppAs determined to date, we report the crystal structure of archaeal OppA from Thermococcus kodakaraensis (TkOppA) at 2.3 Å resolution by the single‐wavelength anomalous dispersion method. TkOppA consists of three domains similarly to bacterial OppAs, and the inserted regions not present in bacterial OppAs are at the periphery of the core region. An endogenous pentapeptide was bound in the pocket of domains I and III of TkOppA by hydrogen bonds of main‐chain atoms of the peptide and hydrophobic interactions. No hydrogen bonds of side‐chain atoms of the peptide were observed; thus, TkOppA may have low peptide selectivity but some preference for residues 2 and 3. TkOppA has a relatively large pocket and can bind a nonapeptide; therefore, it is suitable for the binding of large peptides similarly to OppAs of Gram‐positive bacteria.

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Section: Resultsmentioning

confidence: 99%

Section: Resultsmentioning

confidence: 99%

“…The induced cells were centrifuged and stored at −80°C. Using the cells, the SeMet derivative TkOppA_4mut was purified with the same protocol as described previously 28 …”

Section: Methodsmentioning

confidence: 99%

Section: Preparation Of Tkoppamentioning

confidence: 99%

Section: Structure Determination Of Tkoppamentioning

confidence: 99%

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Structural basis for peptide recognition by archaeal oligopeptide permease A

et al. 2022

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Horizontal Gene Transfer in Archaea—From Mechanisms to Genome Evolution

Gophna

Altman-Price

2022

Annu. Rev. Microbiol.

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Archaea remains the least-studied and least-characterized domain of life despite its significance not just to the ecology of our planet but also to the evolution of eukaryotes. It is therefore unsurprising that research into horizontal gene transfer (HGT) in archaea has lagged behind that of bacteria. Indeed, several archaeal lineages may owe their very existence to large-scale HGT events, and thus understanding both the molecular mechanisms and the evolutionary impact of HGT in archaea is highly important. Furthermore, some mechanisms of gene exchange, such as plasmids that transmit themselves via membrane vesicles and the formation of cytoplasmic bridges that allows transfer of both chromosomal and plasmid DNA, may be archaea specific. This review summarizes what we know about HGT in archaea, and the barriers that restrict it, highlighting exciting recent discoveries and pointing out opportunities for future research. Expected final online publication date for the Annual Review of Microbiology, Volume 76 is September 2022. Please see http://www.annualreviews.org/page/journal/pubdates for revised estimates.

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Preparation, Crystallization, and X-ray Data Collection of Archaeal Oligopeptide Permease A

Cited by 2 publications

References 35 publications

Structural basis for peptide recognition by archaeal oligopeptide permease A

Structural basis for peptide recognition by archaeal oligopeptide permease A

Horizontal Gene Transfer in Archaea—From Mechanisms to Genome Evolution

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