TY - JOUR
T1 - Projection Structure of DtpD (YbgH), a Prokaryotic Member of the Peptide Transporter Family
AU - Casagrande, Fabio
AU - Harder, Daniel
AU - Schenk, Andreas
AU - Meury, Marcel
AU - Ucurum, Zohre
AU - Engel, Andreas
AU - Weitz, Dietmar
AU - Daniel, Hannelore
AU - Fotiadis, Dimitrios
N1 - Funding Information:
This work was supported by the Swiss National Foundation for Scientific Research (Grant 31003A_125150/1 to D.F.), the EC Project Grant 502802 EUGINDAT (to D.F. and H.D.), the Bern University Research Foundation (to D.F. and Matthias Hediger), and the Novartis Foundation (Grant 08A10 to D.F.). We are very grateful to Manuel Palacin (University of Barcelona, Spain) for kindly providing the expression vectors of SteT and AdiC and to Heinz Gross and Peter Tittmann (ETH Zürich, Switzerland) for the unidirectional metal shadowing of tubular DtpD crystals.
PY - 2009/12/11
Y1 - 2009/12/11
N2 - Cellular uptake of di- and tripeptides has been characterized in numerous organisms, and various transporters have been identified. In contrast, structural information on peptide transporters is very sparse. Here, we have cloned, overexpressed, purified, and biochemically characterized DtpD (YbgH) from Escherichia coli, a prokaryotic member of the peptide transporter family. Its homologues in mammals, PEPT1 (SLC15A1) and PEPT2 (SLC15A2), not only transport peptides but also are of relevance for uptake of drugs as they accept a large spectrum of peptidomimetics such as β-lactam antibiotics, antivirals, peptidase inhibitors, and others as substrates. Uptake experiments indicated that DtpD functions as a canonical peptide transporter and is, therefore, a valid model for structural studies of this family of proteins. Blue native polyacrylamide gel electrophoresis, gel filtration, and transmission electron microscopy of single-DtpD particles suggest that the transporter exists in a monomeric form when solubilized in detergent. Two-dimensional crystallization of DtpD yielded first tubular crystals that allowed the determination of a projection structure at better than 19 Å resolution. This structure of DtpD represents the first structural view of a member of the peptide transporter family.
AB - Cellular uptake of di- and tripeptides has been characterized in numerous organisms, and various transporters have been identified. In contrast, structural information on peptide transporters is very sparse. Here, we have cloned, overexpressed, purified, and biochemically characterized DtpD (YbgH) from Escherichia coli, a prokaryotic member of the peptide transporter family. Its homologues in mammals, PEPT1 (SLC15A1) and PEPT2 (SLC15A2), not only transport peptides but also are of relevance for uptake of drugs as they accept a large spectrum of peptidomimetics such as β-lactam antibiotics, antivirals, peptidase inhibitors, and others as substrates. Uptake experiments indicated that DtpD functions as a canonical peptide transporter and is, therefore, a valid model for structural studies of this family of proteins. Blue native polyacrylamide gel electrophoresis, gel filtration, and transmission electron microscopy of single-DtpD particles suggest that the transporter exists in a monomeric form when solubilized in detergent. Two-dimensional crystallization of DtpD yielded first tubular crystals that allowed the determination of a projection structure at better than 19 Å resolution. This structure of DtpD represents the first structural view of a member of the peptide transporter family.
KW - membrane protein
KW - peptide transport protein
KW - structure
KW - transmission electron microscopy
KW - two-dimensional crystal
UR - http://www.scopus.com/inward/record.url?scp=70449524304&partnerID=8YFLogxK
U2 - 10.1016/j.jmb.2009.09.048
DO - 10.1016/j.jmb.2009.09.048
M3 - Article
C2 - 19782088
AN - SCOPUS:70449524304
SN - 0022-2836
VL - 394
SP - 708
EP - 717
JO - Journal of Molecular Biology
JF - Journal of Molecular Biology
IS - 4
ER -