+ Site Statistics
+ Search Articles
+ PDF Full Text Service
How our service works
Request PDF Full Text
+ Follow Us
Follow on Facebook
Follow on Twitter
Follow on LinkedIn
+ Subscribe to Site Feeds
Most Shared
PDF Full Text
+ Translate
+ Recently Requested

Artificial transmembrane segments. Requirements for stop transfer and polypeptide orientation



Artificial transmembrane segments. Requirements for stop transfer and polypeptide orientation



Journal of Biological Chemistry 270(23): 14115-14122



Transmembrane segments of proteins generally consist of a long stretch of hydrophobic amino acids, which can function to initiate membrane insertion (start-stop sequences), initiate translocation (signal-anchor sequences), or stop further translocation of the following polypeptide chain (stop-transfer sequences). In this study, we have taken Escherichia coli alkaline phosphatase, a transported and water-soluble protein, and examined the requirements for converting it into a transmembrane protein with particular orientation. Since the wild type enzyme is transported, there is no predisposition against membrane translocation, yet it is not a membrane protein, so it does not possess any intrinsic membrane topogenic preferences. A series of potential transmembrane segments was introduced into an internal position of the enzyme to test the ability of each to initiate translocation, stop translocation, and adopt a particular orientation. For this purpose, cassette mutagenesis was used to incorporate new structural segments composed of polymers of alanines and leucines. The threshold value of hydrophobicity required to function as a stop-transfer sequence was determined. For a transmembrane segment of typical length (21 residues), this value is equivalent to the hydrophobicity of 16 alanines and 5 leucines. Interestingly, much shorter segments will also suffice to stop translocation, but these must be composed of more highly hydrophobic residues (e.g. 11 leucines). When the wild type amino-terminal signal peptide is deleted or made dysfunctional, sufficiently hydrophobic internal segments can initiate translocation of the following polypeptide and function as a signal anchor. Furthermore, in so doing, the orientation of the protein is changed from N(out)-C(in) to N(in)-C(out).

Please choose payment method:






(PDF emailed within 1 workday: $29.90)

Accession: 008189052

Download citation: RISBibTeXText

PMID: 7775472


Related references

Forced transmembrane orientation of hydrophilic polypeptide segments in multispanning membrane proteins. Molecular Cell 2(4): 495-503, 1998

Negatively charged residues in the IgM stop-transfer effector sequence regulate transmembrane polypeptide integration. Journal of Biological Chemistry 274(47): 33661-33670, 1999

A stop transfer sequence confers predictable transmembrane orientation to a previously secreted protein in cell-free systems. Cell 34(3): 759-766, 1983

Transmembrane protein insertion orientation in yeast depends on the charge difference across transmembrane segments, their total hydrophobicity, and its distribution. Journal of Biological Chemistry 273(38): 24963-24971, 1998

Membrane orientation of transmembrane segments 11 and 12 of MDR- and non-MDR-associated P-glycoproteins. Biochimica et Biophysica Acta 1153(2): 191-202, 1993

Getting greasy: How transmembrane polypeptide segments integrate into the lipid bilayer. Molecular Microbiology 24(2): 249-253, 1997

Membrane orientation of carboxyl-terminal half P-glycoprotein: topogenesis of transmembrane segments. European Journal of Cell Biology 78(9): 624-631, 1999

Helical structure and packing orientation of the four transmembrane segments in the voltage-sensing domain of a K+ channel. Biophysical Journal 78(1 Part 2): 97A, 2000

Structure and packing orientation of transmembrane segments in voltage-dependent channels: lessons from perturbation analysis. The Journal of General Physiology 115(1): 32, 2000

Structure and packing orientation of transmembrane segments in voltage-dependent channels. Lessons from perturbation analysis. Journal of General Physiology 115(1): 29-32, 2000

kPROT: a knowledge-based scale for the propensity of residue orientation in transmembrane segments. Application to membrane protein structure prediction. Journal of Molecular Biology 294(4): 921-935, 1999

A heptad motif of leucine residues found in membrane proteins can drive self-assembly of artificial transmembrane segments. Journal of Biological Chemistry 274(14): 9265-9270, 1999

Type I Interferon Signaling Is Decoupled from Specific Receptor Orientation through Lenient Requirements of the Transmembrane Domain. Journal of Biological Chemistry 291(7): 3371-3384, 2016