Use of amide <sup>1</sup>H‐nmr titration shifts for studies of polypeptide conformation

Bundi, Arno; Wüthrich, Kurt

doi:10.1002/bip.1979.360180207

Cited by 220 publications

(247 citation statements)

References 18 publications

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“…In agreement with earlier observations in different peptides and proteins [14,15], this implies that there is hydrogen bonding between the y-carboxylic acid group and the amide proton of Glu-22. The peptide analogue Arg-Val-Ala-(octahydr0)-Trp-Leu was studied to determine the influence of the ring current field of Trp-23 on the high-field shifts of the resonances of Val-21 (Table 1).…”

Section: Resultssupporting

confidence: 93%

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Characterisation of a Local Structure in the Synthetic Parathyroid Hormone Fragment 1-34 by 1H Nuclear-Magnetic-Resonance Techniques

Bundi¹,

Andreatta

Wüthrich³

1978

Eur J Biochem

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Previous studies had shown that the molecular conformation of the synthetic human parathyroid hormone fragment 1 -34 in dilute aqueous solution contained a local non-random structure formed by the four consecutive residues -Val-21 -Gln-22 -Trp-23 -Leu-24 -. This paper gives a detailed description of this local spatial structure obtained from high resolution 'H NMR studies at 360 MHz of several peptide analogs of the partial sequence 20-24. The most important spectral parameters were high-field shifts of the a and y protons of Val-21, the spin-spin coupling constants related to the rotamer populations of the side-chains of Val-21 and Trp-23, and pH titration shifts of the amide proton resonances. It was found that the backbone fragment 20-24 is so arranged that the side-chain of Val-21 is located next to the indole ring plane of Trp-23; evidence is presented that this non-random structure is mainly stabilized by hydrophobic interactions between the side-chains of Val-21 and Trp-23. The thermal population of the observed molecular structure at room temperature was estimated from the nuclear magnetic resonance data to be approximately 20%.In a previous paper it was shown that the synthetic human parathyroid hormone fragment 1 -34 exists in aqueous solution predominantly in a flexible extended form, where, however, a local non-random structure was also clearly manifested in the ' H nuclear magnetic resonance (NMR) parameters [l]. This non-random conformation was then found to involve the amino acid residues -Val-21 -Gln-22-Trp-23 -Leu-24 -, and it was demonstrated that the same structure was preserved in the pentapeptide Arg-Val-Gln-Trp-Leu[l]. Our interest in this structure was further enhanced, since the tetrapeptide sequence 21 -24 in the parathyroid hormone corresponds to the tetrapeptide sequence 23 -26 in glucagon [2,3]. The present paper describes a detailed investigation of the spatial arrangement of the tetrapeptide segment 21 -24 in the parathyroid hormone 1 -34.The strategy used for this study was to compare the spatial structures of different synthetic peptide analogs by high-resolution 'H NMR techniques. On the basis of the requirements of compatibility with all the NMR data and with the criteria of closest interAbhreviations. NMR, nuclear magnetic resonance; parathyroid hormone 1 -34, synthetic N-terminal fragment 1 -34 of the human parathyroid hormone. atomic distances, a molecular model for the tripeptide segment -Val-21 -Gln-22 -Trp-23 -in the parathyroid hormone 1 -34 was constructed. With the use of the atomic coordinates obtained from this model, the thermal population of the local spatial structure was estimated in a quantitative analysis of the NMR data. MATERIALS AND METHODS Peptidr SynthesisThe synthesis of parathyroid hormone 1-34 according to the sequence determined by Brewer et al. [4] and of several partial sequences thereof was described previously [ 5 ] . This concerns in particular the partial sequences Met-1

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Section: Resultssupporting

confidence: 93%

“…The distinction between the states I1 and I11 is possible only when the two methylene protons Ha* and HPB are stereospecifically assigned. A systematic study of the linear tetrapeptides Gly-Gly-X-Ala provided a criterion for a stereospecific assignment of the b methylene protons of aromatic side-chains [8]. Accordingly, HPA (Fig.…”

Section: Resultsmentioning

confidence: 99%

Characterisation of a Local Structure in the Synthetic Parathyroid Hormone Fragment 1-34 by 1H Nuclear-Magnetic-Resonance Techniques

Bundi¹,

Andreatta

Wüthrich³

1978

Eur J Biochem

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“…Stretches of consecutive amide-amide NOEs (dNN), indicative for formation of local structures, were absent as were intermediate or long range NOE contacts. All of the experimentally determined coupling constants between amide and ␣-protons showed values very close to those reported for random coil peptides (31) (Fig. 3).…”

Section: ␤N-(1-62) Interacts With the ␣-Subunit Of Kv 11 Channels Insupporting

confidence: 81%

NMR Structure and Functional Characteristics of the Hydrophilic N Terminus of the Potassium Channel β-Subunit Kvβ1.1

Wissmann

Baukrowitz

Kalbacher

et al. 1999

Journal of Biological Chemistry

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Rapid N-type inactivation of voltage-dependent potassium (Kv) channels controls membrane excitability and signal propagation in central neurons and is mediated by protein domains (inactivation gates) occluding the open channel pore from the cytoplasmic side. Inactivation domains (ID) are donated either by the pore-forming ␣-subunit or certain auxiliary ␤-subunits. Upon coexpression, Kv␤1.1 was found to endow non-inactivating members of the Kv1␣ family with fast inactivation via its unique N terminus. Here we investigated structure and functional properties of the Kv␤1.1 N terminus (amino acids 1-62, ␤N-(1-62)) using NMR spectroscopy and patch clamp recordings. ␤N-(1-62) showed all hallmarks of N-type inactivation: it inactivated non-inactivating Kv1.1 channels when applied to the cytoplasmic side as a synthetic peptide, and its interaction with the ␣-subunit was competed with tetraethylammonium and displayed an affinity in the lower micromolar range. In aequous and physiological salt solution, ␤N-(1-62) showed no well defined three-dimensional structure, it rather existed in a fast equilibrium of multiple weakly structured states. These structural and functional properties of ␤N-(1-62) closely resemble those of the "unstructured" ID from Shaker B, but differ markedly from those of the compactly folded ID of the Kv3.4 ␣-subunit.Fast N-type inactivation of voltage-gated potassium (Kv) 1 channels shapes the action potential, governs the firing rate (spiking), and controls signal propagation in central neurons (1). Biophysically, N-type inactivation has long served as the model for gating transitions in ion channels and is realized by a "ball plug-in" mechanism. In this mechanism a protein domain termed "inactivation gate" or "inactivation ball" binds to its receptor at the inner vestibule of the open channel and thereby occludes the ion pathway (2-5). Such inactivation gates have been localized in the N terminus of various Kv␣ subunits and were shown to be functional entities, i.e. they conferred rapid inactivation to "ball-less" Kv␣ subunits when applied to the cytoplasmic side of the channels as synthetic peptides (5-8). Identical to protein-harbored inactivation domains, the synthetic gates interacted with channels in the open state, blocked the pore with low voltage dependence, and were competed with the channel blocker tetraethylammonium (TEA) (9 -11). Recently, the structures of the inactivation domains (ID) from Kv1.4 and Kv3.4 were determined with NMR spectroscopy. Both IDs were found to exhibit well defined and compact folding in aequous solution (12). In contrast, the ID from Shaker B showed no unique, folded structure (13,14).Besides with Kv␣ subunits owning an N-terminal ID, fast inactivation was observed for a subset of non-inactivating Kv␣1 channels when coexpressed with certain ␤-subunits (15-17). These auxiliary subunits constitute a family of cytoplasmic proteins (subdivided into subfamilies Kv␤1, -2, and -3) that are made up of two distinct regions: a highly conserved core region that shows homolo...

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“…The two doublets at 6.62 ppm and 7.03 ppm are symmetrically disposed about the skew diagonal and thus assigned to H3 and H5, and H2 and H6 of the tyrosine residue. The upfield doublet of the Tyr spin system is assigned to H3 and H5 and, the downfield doublet to H6 and H2 [35]. A triplet at 6.88 ppm is symmetrically related to a resonance at 7.09 ppm which is I the skew diagonal, due to the overlap of the aromatic signals.…”

Section: Aromaric Residuesmentioning

confidence: 99%

Preliminary assignments of the aromatic and some methyl group resonances of the ¹H‐NMR spectrum of the oxidized form of uteroglobin

Jamin¹,

Roy

Fridlansky

et al. 1989

European Journal of Biochemistry

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Two‐dimensional NMR methods have been used to assign aromatic and methyl group resonances in the 1H‐NMR spectrum of oxidized uteroglobin. Assignments to specific amino acids are based on X‐ray‐determined structures of two crystal forms (C2221 and P21) and on an energy‐minimized X‐ray structure of the C2221 form of uteroglobin. These preliminary assignments are sufficient to probe the interaction of oxidized uteroglobin with progesterone in solution. The protein global structure is unmodified but some direct or indirect conformational changes are induced in the H1H4(H1′H4′) pockets and close to Phe28 by progesterone.

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Use of amide ¹H‐nmr titration shifts for studies of polypeptide conformation

Cited by 220 publications

References 18 publications

Characterisation of a Local Structure in the Synthetic Parathyroid Hormone Fragment 1-34 by 1H Nuclear-Magnetic-Resonance Techniques

Characterisation of a Local Structure in the Synthetic Parathyroid Hormone Fragment 1-34 by 1H Nuclear-Magnetic-Resonance Techniques

NMR Structure and Functional Characteristics of the Hydrophilic N Terminus of the Potassium Channel β-Subunit Kvβ1.1

Preliminary assignments of the aromatic and some methyl group resonances of the ¹H‐NMR spectrum of the oxidized form of uteroglobin

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Use of amide 1H‐nmr titration shifts for studies of polypeptide conformation

Cited by 220 publications

References 18 publications

Characterisation of a Local Structure in the Synthetic Parathyroid Hormone Fragment 1-34 by 1H Nuclear-Magnetic-Resonance Techniques

Characterisation of a Local Structure in the Synthetic Parathyroid Hormone Fragment 1-34 by 1H Nuclear-Magnetic-Resonance Techniques

NMR Structure and Functional Characteristics of the Hydrophilic N Terminus of the Potassium Channel β-Subunit Kvβ1.1

Preliminary assignments of the aromatic and some methyl group resonances of the 1H‐NMR spectrum of the oxidized form of uteroglobin

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Use of amide ¹H‐nmr titration shifts for studies of polypeptide conformation

Preliminary assignments of the aromatic and some methyl group resonances of the ¹H‐NMR spectrum of the oxidized form of uteroglobin