Gpi

transamidase

NH3+ protein ER lumen

Mature GPI-linked protein

NH3+ protein ER lumen

Mature GPI-linked protein

▲ FIGURE 16-14 GPI-anchored proteins. (a) Structure of a glycosylphosphatidylinositol (GPI) from yeast. The hydrophobic portion of the molecule is composed of fatty acyl chains, whereas the polar (hydrophilic) portion of the molecule is composed of carbohydrate residues and phosphate groups. In other organisms, both the length of the acyl chains and the carbohydrate moieties may vary somewhat from the structure shown. (b) Formation of GPI-anchored proteins in the ER membrane. The protein is synthesized and initially inserted into the ER membrane as shown in Figure 16-11. A specific transamidase simultaneously cleaves the precursor protein within the exoplasmic-facing domain, near the stop-transfer anchor sequence (red), and transfers the carboxyl group of the new C-terminus to the terminal amino group of a preformed GPI anchor. [See C. Abeijon and C. B. Hirschberg, 1992, Trends Biochem. Sci. 17:32, and K. Kodukula et al., 1992, Proc. Nat'l. Acad. Sci. USA 89:4982.]

stop-transfer anchor sequence and transfers the remainder of the protein to a preformed GPI anchor in the membrane (Figure 16-14b).

Why change one type of membrane anchor for another? Attachment of the GPI anchor, which results in removal of the cytosol-facing hydrophilic domain from the protein, can have several consequences. Proteins with GPI anchors, for example, can diffuse in the plane of the phospholipid bilayer membrane. In contrast, many proteins anchored by membrane-spanning a helices are immobilized in the membrane because their cytosol-facing segments interact with

The Topology of a Membrane Protein Often Can Be Deduced from Its Sequence

As we have seen, various topogenic sequences in integral membrane proteins synthesized on the ER govern interaction of the nascent chain with the translocon. When scientists begin to study a protein of unknown function, the identification of topogenic sequences within the corresponding gene sequence can provide important clues about the protein's topological class and function. Suppose, for example, that the gene for a protein known to be required for a cell-to-cell signaling pathway contains nucleotide sequences that encode an apparent N-terminal signal sequence and an internal hy-drophobic sequence. These findings would suggest that the protein is a type I integral membrane protein and therefore may be a cell-surface receptor for an extracellular ligand.

Identification of topogenic sequences requires a way to scan sequence databases for segments that are sufficiently hy-drophobic to be either a signal sequence or a transmembrane anchor sequence. Topogenic sequences can often be identified with the aid of computer programs that generate a hydropathy profile for the protein of interest. The first step is to assign a value known as the hydropathic index to each amino acid in the protein. By convention, hydrophobic amino acids are assigned positive values, and hydrophilic amino acids negative values. Although different scales for the hydropathic index exist, all assign the most positive values to amino acids with side chains made up of mostly hydrocarbon residues (e.g., phenylalanine and methionine) and the most negative values to charged amino acids (e.g., arginine and aspartate). The second step is to identify longer segments of sufficient overall hydrophobicity to be N-terminal signal sequences or internal stop-transfer sequences and signal-anchor sequences. To accomplish this, the total hydropathic index for each successive sliding "window" of 20 consecutive amino acids is calculated along the entire length of the protein. Plots of these calculated values against position in the amino acid sequence yield a hydropathy profile.

Figure 16-15 shows the hydropathy profiles for three different membrane proteins. The prominent peaks in such plots identify probable topogenic sequences, as well as their position and approximate length. For example, the hydropathy profile of the human growth hormone receptor reveals the presence of both a hydrophobic signal sequence at the extreme N-terminus of the protein and an internal hydropho-bic stop-transfer sequence (see Figure 16-15a). On the basis

NH3+

NH3+

(a) Human growth hormone receptor (type I)

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