scholarly journals 1H-n.m.r. study of the solution properties and secondary structure of neurotoxin III from the sea anemone Anemonia sulcata

1993 ◽  
Vol 293 (2) ◽  
pp. 545-551 ◽  
Author(s):  
R S Norton ◽  
K Cross ◽  
V Braach-Maksvytis ◽  
E Wachter
Keyword(s):  
Secondary Structure ◽  
Electrostatic Interactions ◽  
Hydrophobic Interactions ◽  
Nuclear Overhauser Effect ◽  
Resonance Assignments ◽  
Alpha Helix ◽  
Sea Anemone ◽  
Coupling Constants ◽  
Solution Properties ◽  
Anemonia Sulcata

The solution properties, secondary structure and global fold of the 27-residue polypeptide neurotoxin III (ATX III), from the sea anemone Anemonia sulcata, have been investigated using high-resolution 1H-n.m.r. spectroscopy. Studies of the concentration dependence of the n.m.r. spectrum indicate that the molecule self-associates in the millimolar concentration range useable for n.m.r. analysis, the association being less pronounced at acidic pH values. The dependence on pH of association implies that electrostatic interactions play a role in this process, while the significant concentration-dependent shifts of the aromatic resonances of Tyr-7 and Trp-13 indicate that hydrophobic interactions also contribute. Individual pKa values have been determined for most ionizable groups in the molecule. Sequence-specific resonance assignments were obtained for all protons using a range of two-dimensional homonuclear-correlated and nuclear-Overhauser-effect (nOe) spectra. The secondary structure of the polypeptide was identified from sequential (i, i+1) and medium-range (i, i+2/3/4) nOe connectivities, NH to C alpha H coupling constants, C alpha H chemical shifts, and the location of slowly exchanging backbone-amide protons. ATX III contains no regular alpha-helix or beta-sheet, consisting instead of a network of reverse turns. nOe connectivities between half-cystine residues are consistent with the disulphide pairings 3-17, 4-11 and 6-22. ATX III has a well-defined structure and appears to lack the disordered loop which, in the longer sea anemone toxins (46-49 residues), may be part of the receptor-binding surface.

scholarly journals Multiple native-like conformations trapped via self-association-induced hydrophobic collapse of the 33-residue β-sheet domain from platelet factor 4

1995 ◽  
Vol 306 (2) ◽  
pp. 407-419 ◽  
Author(s):  
E Ilyina ◽  
K H Mayo
Keyword(s):  
Electrostatic Interactions ◽  
Nuclear Overhauser Effect ◽  
Alpha Helix ◽  
Platelet Factor 4 ◽  
Random Coil ◽  
Beta Sheet ◽  
Platelet Factor ◽  
Hydrophobic Collapse ◽  
Low Dielectric ◽  
Self Association

Native platelet factor 4 (PF4) (70 residues) has a hydrophobic three-stranded anti-parallel beta-sheet domain on to which is folded an amphipathic C-terminal alpha-helix and an aperiodic N-terminal domain. The 33-amino acid beta-sheet domain from PF4 (residues 23-55) has been synthesized and studied by c.d. and n.m.r. At 10 degrees C and low concentration, peptide 23-55 appears to exist in aqueous solution in a random-coil distribution of highly flexible conformational states. Some preferred conformation, however, is observed, particularly within a relatively stable chain reversal from Leu-45 to Arg-49. As the peptide concentration and/or temperature is increased, a new conformational state(s) appears and intensifies as slowly exchanging (600 MHz 1H-n.m.r. chemical-shift time scale) random-coil resonances disappear. Hill plots of the concentration-dependence indicated mostly tetramer formation as found in native PF4. Although apparent resonance linewidths in aggregate state(s) are of the order of 100 Hz, sequence-specific assignments for most resonances could be made. N.m.r./nuclear Overhauser effect structural analysis indicates the formation of multiple native-like anti-parallel beta-sheet conformations, kinetically trapped via subunit-association-induced hydrophobic collapse and stabilized by low-dielectric electrostatic interactions among/between Gly-28 and Lys-50 in opposing subunits. Results are discussed in terms of protein folding.

Sequence Specificity Despite Intrinsic Disorder: How a Disease-Associated Val/Met Polymorphism Shifts Tertiary Interactions in a Long Disordered Protein

2019 ◽  
Author(s):  
Ruchi Lohia ◽  
Reza Salari ◽  
Grace Brannigan
Keyword(s):  
Secondary Structure ◽  
Electrostatic Interactions ◽  
Intrinsically Disordered Proteins ◽  
Disordered Proteins ◽  
Sequence Specificity ◽  
Intrinsically Disordered ◽  
Specific Effects ◽  
Heterogeneous Polymer ◽  
Non Local ◽  
Dynamics Simulations

<div>The role of electrostatic interactions and mutations that change charge states in intrinsically disordered proteins (IDPs) is well-established, but many disease-associated mutations in IDPs are charge-neutral. The Val66Met single nucleotide polymorphism (SNP) encodes a hydrophobic-to-hydrophobic mutation at the midpoint of the prodomain of precursor brain-derived neurotrophic factor (BDNF), one of the earliest SNPs to be associated with neuropsychiatric disorders, for which the underlying molecular mechanism is unknown. Here we report on over 250 μs of fully-atomistic, explicit solvent, temperature replica exchange molecular dynamics simulations of the 91 residue BDNF prodomain, for both the V66 and M66 sequence.</div><div>The simulations were able to correctly reproduce the location of both local and non-local secondary changes due to the Val66Met mutation when compared with NMR spectroscopy. We find that the local structure change is mediated via entropic and sequence specific effects. We show that the highly disordered prodomain can be meaningfully divided into domains based on sequence alone. Monte Carlo simulations of a self-excluding heterogeneous polymer, with monomers representing each domain, suggest the sequence would be effectively segmented by the long, highly disordered polyampholyte near the sequence midpoint. This is qualitatively consistent with observed interdomain contacts within the BDNF prodomain, although contacts between the two segments are enriched relative to the self-excluding polymer. The Val66Met mutation increases interactions across the boundary between the two segments, due in part to a specific Met-Met interaction with a Methionine in the other segment. This effect propagates to cause the non-local change in secondary structure around the second methionine, previously observed in NMR. The effect is not mediated simply via changes in inter-domain contacts but is also dependent on secondary structure formation around residue 66, indicating a mechanism for secondary structure coupling in disordered proteins. </div>

1H, 15N and 13C resonance assignments and secondary structure of PulG, the major pseudopilin from Klebsiella oxytoca type 2 secretion system

2017 ◽  
Vol 11 (2) ◽  
pp. 155-158 ◽  
Author(s):  
Aracelys López-Castilla ◽  
Bruno Vitorge ◽  
Léa Khoury ◽  
Nelly Morellet ◽  
Olivera Francetic ◽  
...  
Keyword(s):  
Secondary Structure ◽  
Klebsiella Oxytoca ◽  
Resonance Assignments ◽  
Secretion System ◽  
Type 2 Secretion

scholarly journals Characterization of porcine osteonectin extracted from foetal calvariae

1988 ◽  
Vol 253 (1) ◽  
pp. 139-151 ◽  
Author(s):  
C Domenicucci ◽  
H A Goldberg ◽  
T Hofmann ◽  
D Isenman ◽  
S Wasi ◽  
...  
Keyword(s):  
Amino Acids ◽  
Secondary Structure ◽  
Type I Collagen ◽  
Culture Shock ◽  
Gel Filtration ◽  
Alpha Helix ◽  
Type I ◽  
Homologous Proteins ◽  
Compact Structure ◽  
Significant Difference

Osteonectin, extracted from foetal porcine calvariae with 0.5 M-EDTA, was purified to homogeneity by using gel filtration and polyanion anion-exchange fast protein liquid chromatography under dissociative conditions without the need of reducing agents. The purified protein migrated with an Mr of 40,300 on SDS/polyacrylamide gels and was similar to bovine osteonectin in both amino acid composition and in its ability to bind to hydroxyapatite in the presence of 4 M-guanidinium hydrochloride (GdmCl). However, unlike the bovine protein, porcine osteonectin did not bind selectively to hydroxyapatite when EDTA tissue extracts were used. In addition, purified porcine osteonectin did not show any apparent affinity for either native or denatured type I collagen, but did bind to serum albumin. Primary sequence analysis revealed an N-terminal alanine residue, with approximately one-half of the subsequent 35 residues identified as small hydrophobic amino acids and one-quarter as acidic amino acids. The only significant difference between the N-terminal sequences of the bovine and porcine proteins was the deletion of the tripeptide Val-Ala-Glu in porcine osteonectin. In contrast with bovine osteonectin, far-u.v.c.d. of porcine osteonectin revealed considerable secondary structure, of which 27% was alpha-helix and 39% was beta-sheet. Cleavage of the molecule with CNBr under non-reducing conditions generated five fragments, of which two major fragments (Mr 27,900 and 12,400) stained blue with Stains All, a reagent that stains sialic-acid-rich proteins/phosphate-containing proteins and/or Ca2+-binding proteins blue while staining other proteins pink. The 12,400-Mr fragment bound 45Ca2+ selectively, indicating a Ca2+-binding site in this part of the molecule. The 27,900-Mr fragment did not bind Ca2+, and since biosynthetic studies with 32PO4(3-) did not show phosphorylation of porcine osteonectin, this fragment is likely to be highly acidic. The incomplete cleavage of the molecule with CNBr and the ability of the molecule to regain its secondary structure after exposure to 7 M-urea are features consistent with the molecule having a compact structure that is stabilized by numerous disulphide bridges. The chemical and binding properties of porcine osteonectin are closely similar to the recently described ‘culture shock’, SPARC and BM-40 proteins, indicating that these are homologous proteins.

1H, 13C, and 15N NMR Resonance Assignments, Secondary Structure, and Backbone Topology of a Variant of Human Interleukin-3

Biochemistry ◽  
1995 ◽  
Vol 34 (19) ◽  
pp. 6540-6551 ◽  
Author(s):  
Yiqing Feng ◽  
Barbara K. Klein ◽  
Linh Vu ◽  
Serdar Aykent ◽  
Charles A. McWherter
Keyword(s):  
Secondary Structure ◽  
Resonance Assignments ◽  
Nmr Resonance Assignments ◽  
15N Nmr ◽  
Interleukin 3

15N, 13C and 1H resonance assignments and secondary structure determination of a variable heavy domain of a heavy chain antibody

2013 ◽  
Vol 8 (1) ◽  
pp. 113-116 ◽  
Author(s):  
Christine E. Prosser ◽  
Lorna C. Waters ◽  
Frederick W. Muskett ◽  
Vaclav Veverka ◽  
Philip W. Addis ◽  
...  
Keyword(s):  
Secondary Structure ◽  
Heavy Chain ◽  
Structure Determination ◽  
Resonance Assignments ◽  
Variable Heavy ◽  
Heavy Chain Antibody

scholarly journals Characterization of Virus Adsorption by Using DEAE-Sepharose and Octyl-Sepharose

2002 ◽  
Vol 68 (8) ◽  
pp. 3965-3968 ◽  
Author(s):  
Patricia A. Shields ◽  
Samuel R. Farrah
Keyword(s):  
Sodium Chloride ◽  
Electrostatic Interactions ◽  
Hydrophobic Interactions ◽  
Chloride Concentration ◽  
Sodium Chloride Concentration ◽  
Virus Adsorption ◽  
Nacl Concentration

ABSTRACT Viruses were characterized by their adsorption to DEAE-Sepharose or by their elution from octyl-Sepharose by using buffered solutions of sodium chloride with different ionic strengths. Viruses whose adsorption to DEAE-Sepharose was reduced most rapidly by an increase in the sodium chloride concentration were considered to have the weakest electrostatic interactions with the solids; these viruses included MS2, E1, and φX174. Viruses whose adsorption to DEAE-Sepharose was reduced least rapidly were considered to have the strongest electrostatic interactions with the column; these viruses included P1, T4, T2, and E5. All of the viruses studied adsorbed to octyl-Sepharose in the presence of 4 M NaCl. Viruses that were eluted most rapidly following a decrease in the concentration of NaCl were considered to have the weakest hydrophobic interactions with the column; these viruses included φX174, CB4, and E1. Viruses that were eluted least rapidly from the columns after the NaCl concentration was decreased were considered to have the strongest hydrophobic interactions with the column; these viruses included f2, MS2, and E5.

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