Refinement of the crystal structure of wheat germ agglutinin isolectin 2 at 1.8 Å resolution

Wright, Christopher S.

doi:10.1016/0022-2836(87)90678-4

Cited by 90 publications

(23 citation statements)

References 44 publications

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“…Extensive x-ray crystallographic and sequence analyses have revealed that mature WGA is a homodimeric protein composed of 18-kD subunits. Each subunit is composed of four homologous domains, each of which consists of a tightly folded core stabilized by four disulfide bonds (Wright, 1987). Examination of the WGA crystal structure does not reveal any regions that extend from the surface.…”

Section: Mechanisms Of Sorting To the Vacuolementioning

confidence: 99%

Determination of the functional elements within the vacuolar targeting signal of barley lectin.

et al. 1993

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We have previously demonstrated that the carboxyl-terminal propeptide of barley lectin is both necessary and sufficient for protein sorting to the plant vacuole. Specific mutations were constructed to determine which amino acid residues or secondary structural determinants of the carboxyl-terminal propeptide affect proper protein sorting. We have found that no consensus sequence or common structural determinants are required for proper sorting of barley lectin to the vacuole. However, our analysis demonstrated the importance of hydrophobic residues in vacuolar targeting. In addition, at least three exposed amino acid residues are necessary for efficient sorting. Sorting was disrupted by the addition of two glycine residues at the carboxyl-terminal end of the targeting signal or by the translocation of the glycan to the carboxy terminus of the propeptide. These results suggest that some components of the sorting apparatus interact with the carboxy terminus of the propeptide.

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Section: Mechanisms Of Sorting To the Vacuolementioning

confidence: 99%

Determination of the functional elements within the vacuolar targeting signal of barley lectin.

et al. 1993

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“…Concomitant with the formation of an active lectin dimer, the proprotein assumed a conformation in which the highmannose glycan sequestered the propeptide from the aqueous environment, thereby masking the availability of the propeptide for processing (Figure 9, top panel). This predicted protein configuration was predicated on the conformation of the protein (Wright, 1987), the amphipathic characteristic of the propeptide, and the hydrophilic nature of the glycan. In the trans-cisternae of the Golgi complex, the glycan is removed post-translationally from the proprotein in a regulated manner.…”

Section: A Model For the Role Of The Glycan In The Posttranslational mentioning

confidence: 99%

“…The processing model schematically depicts one subunit of a barley lectin dimer adapted from the structure of WGA (Wright, 1987). Each of the highly homologous domains of barley lectin is represented by a circle.…”

Section: Nucleic Acid Analysismentioning

confidence: 99%

“…Nitrocellulose filters were hybridized with 32P randomprimer-labeled (Feinberg and Vogelstein, 1983) BLc3 barley lectin cDNA (Lerner and Raikhel, 1989) as described previously . For gene reconstruction experiments, tobacco genomic DNA was restricted with EcoRl and BLc3 titered at 0.5-copy, 1 .O-copy, 3.O-copy, and 5.0-copy equivalents per tobacco genome (4.8 x 10g bp per haploid genome; Zimmerman and Goldberg, 1977). Gene reconstruction blots were hybridized with a radiolabeled BLc3 insert by the random-primer method (Feinberg and Vogelstein, 1983).…”

Section: Site-directed Mutagenesismentioning

confidence: 99%

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Role of propeptide glycan in post-translational processing and transport of barley lectin to vacuoles in transgenic tobacco.

1990

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Mature barley lectin is a dimeric protein composed of two identical 18-kilodalton polypeptides. The subunits of barley lectin are initially synthesized as glycosylated proproteins, which are post-translationally processed to the mature protein preceding or concomitant with deposition of barley lectin in vacuoles. To investigate the functional role of the glycan in processing and intracellular transport of barley lectin to vacuoles, the sole N-linked glycosylation site residing within the COOH-terminal propeptide of barley lectin was altered by site-directed mutagenesis. cDNA clones encoding wild-type (wt) or glycosylation-minus ( gly-) barley lectin preproproteins were placed under the transcriptional control of the cauliflower mosaic virus 35s promoter and introduced into Nicotiana tabacum cv Wisconsin 38. Barley lectin synthesized from both the wt and g/y-constructs was processed and correctly targeted to vacuoles of tobacco leaves. Localization of barley lectin in vacuoles processed from the nonglycosylated g/yproprotein indicated that the high-mannose glycan of the barley lectin proprotein was not essential for targeting barley lectin to vacuoles. However, pulse-chase labeling experiments demonstrated that the glycosylated wt proprotein and the nonglycosylated g/y-proprotein were differentially processed to the mature protein and transported from the Golgi complex at different rates. These results implicate an indirect functional role for the glycan in post-translational processing and transport of barley lectin to vacuoles.

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“…The 3D structures of a number of these snake venom neurotoxins have been determined using X-ray crystallography and NMR (Bourne et al, 1985;Love and Stroud, 1986;Labhardt et al, 1988;Betzel et al, 1991;Oswald et al, 1991;le Du et al, 1992;Le Goas et al, 1992;Yu et al, 1993;and references therein). These structures display a common folding topology, the so-called toxinagglutinin motif (Drenth et al, 1980), which is also found in snake venom cardiotoxins (Rees et al, 1990;Steinmetz et al, 1988;and references therein) and the domains of WGA (Wright, 1987, and references therein). Harrison (1992) observed that the 10 cysteine residues of CD59 are highly conserved in the snake venom neurotoxins and that their disulfide bonding pattern is similar or identical (the preliminary report of Tomita et al [1991] describes the CD59 disulfide pattern with one ambiguity regarding the bonding of the two adjacent cysteines 63 and 64).…”

mentioning

confidence: 98%

Sequence‐specific

Fletcher¹,

Harrison²,

Lachmann³

et al. 1993

Protein Science

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CD59 is a recently discovered cell-surface glycoprotein that restricts lysis by homologous complement and has limited sequence similarity to snake venom neurotoxins. This paper describes the first results of a two-dimensional NMR study of CD59 prepared from human urine. Nearly complete 'H-NMR assignments were obtained for the 77 amino acid residues and partial assignments for the N-glycan and the glycosylphosphatidylinositol (GPI) anchor. These results together confirm that the C-terminal residue of the mature protein is Asn 77 and that the urine-derived form retains the nonlipid part of the GPI anchor. The data further indicate that the GPI anchor and possibly the N-glycan are structurally inhomogeneous and suggest that the phospholipid present in the intact GPI anchor was removed by phosphatidylinositol-specific phospholipase-D. The folding topology of the protein was determined from NOE enhancements and slowly exchanging backbone amide protons and consists primarily of five extended strands (denoted p l -f l 5 in sequence order), arranged into separate two-stranded (PI and &) and three-stranded (&&) antiparallel P-sheets. The same folding topology is found in all of the snake venom neurotoxins whose structures have been determined. The region between the & and & strands has helical character, a feature that is not present in the neurotoxins but that is seen in the topologically similar wheat germ agglutinin.

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Refinement of the crystal structure of wheat germ agglutinin isolectin 2 at 1.8 Å resolution

Cited by 90 publications

References 44 publications

Determination of the functional elements within the vacuolar targeting signal of barley lectin.

Determination of the functional elements within the vacuolar targeting signal of barley lectin.

Role of propeptide glycan in post-translational processing and transport of barley lectin to vacuoles in transgenic tobacco.

Sequence‐specific

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