HSQC-based Methyl Group Selection via Gradients in Multidimensional NMR Spectroscopy of Proteins

✍ Scribed by Tammo Diercks; Manfred Schwaiger; Horst Kessler

• Book ID: 102594574
• Publisher: Elsevier Science
• Year: 1998
• Tongue: English
• Weight: 186 KB
• Volume: 130
• Category: Article
• ISSN: 1090-7807
• DOI: 10.1006/jmre.1997.1321

✦ Synopsis

A quadruple-quantum filtered HSQC ( QQF-HSQC ) for the and demonstrated in practice on the 14-kDa human nonpancreatic synovial phospholipase A2 ( hnps-PLA2 ) . The constant-time selection of methyl groups which minimizes the line-broadening effects of proton homonuclear couplings is presented. The scheme version is particularly useful for obtaining high-resolution spectra as demonstrated on hnps-PLA2. The applicability of the CT-uses gradients for n -quantum coherence selection and solvent suppression. In contrast to the heteronuclear quadruple-quantum QQF-HSQC module in multidimensional experiments is demonstrated using a 3D CT NOESY-QQF-HSQC spectrum of the 31-coherence ( HQQC ) approach, the QQF-HSQC allows for long constant-time ( CT) evolution, making use of the generally favor-kDa homodimeric IIAMan domain of the mannose transporter of E. coli. ᭧ 1998 Academic Press able relaxation properties of methyl groups. The increase in resolution and concomitant gain in sensitivity is discussed in theory FIG. 1. Pulse sequence of the constant-time quadruple-quantum filtered HSQC (CT-QQF-HSQC) experiment with the relevant coherence-transfer pathways. Narrow and wide bars indicate 90Њ and 180Њ pulses, respectively.

All pulses were applied with phase x if not otherwise indicated. The phase cycles used in this experiment were f 1 Å x, 0x ; f 2 Å 2x, 2(0x) and f rec Å x, 2(0x), x. In the constant-time period the arrow indicates the direction in which the pulse is shifted as t 1 is incremented; the delays were set as follows: D Å 2 ms, CT Å 12.3 ms. Proton decoupling was achieved by a MLEV16 expansion of 180Њ pulses. Frequency discrimination in F 1 was achieved using the echo-antiecho procedure. Pulsed field gradients of relative amplitudes G 1 Å 20, G 2 Å 020, G 3 Å 65 for p-selection and G 1 Å 20, G 2 Å 020, G 3 Å 55 for the n-selection were applied for a duration of 1.0 ms, followed by 0.2 ms recovery delays. All gradient pulses were applied at an angle of 54.7Њ relative to the z coordinate according to the relation G x Å 2rG z . 335