Pro4 prolyl peptide bond isomerization in human galectin-7 modulates the monomer-dimer equilibrum to affect function.

Human galectin-7 (Gal-7; also termed p53-induced gene 1 product) is a multifunctional effector by productive pairing with distinct glycoconjugates and protein counter-receptors in the cytoplasm and nucleus, as well as on the cell surface. Its structural analysis by NMR spectroscopy detected doubling of a set of particular resonances, an indicator of Gal-7 existing in two conformational states in slow exchange on the chemical shift time scale. Structural positioning of this set of amino acids around the P4 residue and loss of this phenomenon in the bioactive P4L mutant indicated cis-trans isomerization at this site.

Lactose binding to human galectin-7 (p53-induced gene 1) induces long-range effects through the protein resulting in increased dimer stability and evidence for positive cooperativity.

The product of p53-induced gene 1 is a member of the galectin family, i.e., galectin-7 (Gal-7).

Lactose binding to galectin-1 modulates structural dynamics, increases conformational entropy, and occurs with apparent negative cooperativity.

Galectins are a family of lectins with a conserved carbohydrate recognition domain that interacts with beta-galactosides. By binding cell surface glycoconjugates, galectin-1 (gal-1) is involved in cell adhesion and migration processes and is an important regulator of tumor angiogenesis. Here, we used heteronuclear NMR spectroscopy and molecular modeling to investigate lactose binding to gal-1 and to derive solution NMR structures of gal-1 in the lactose-bound and unbound states.

A simple method to measure 13CH2 heteronuclear dipolar cross-correlation spectral densities.

Here, we report a method to simultaneously determine CH2 cross-correlation spectral densities and T1 relaxation times in the laboratory and rotating frames. To accomplish this, we have employed an indirect approach that is based on measurement of differences in relaxation rates acquired with and without cross-correlation terms. The new method, which can be employed using multidimensional NMR and standard relaxation pulse sequences, is validated experimentally by investigation of a selectively 13C-enriched hexadecapeptide and the uniformly 13C-enriched immunoglobulin-binding domain of streptococcal protein G (GB1).

Comparison of (13)C(alpha)H and (15)NH backbone dynamics in protein GB1.

This study presents a site-resolved experimental view of backbone C(alpha)H and NH internal motions in the 56-residue immunoglobulin-binding domain of streptococcal protein G, GB1. Using (13)C(alpha)H and (15)NH NMR relaxation data [T(1), T(2), and NOE] acquired at three resonance frequencies ((1)H frequencies of 500, 600, and 800 MHz), spectral density functions were calculated as F(omega) = 2omegaJ(omega) to provide a model-independent way to visualize and analyze internal motional correlation time distributions for backbone groups in GB1. Line broadening in F(omega) curves indicates the presence of nanosecond time scale internal motions (0.

Protein dynamics using frequency-dependent order parameters from analysis of NMR relaxation data.

A novel approach is described to analyze NMR relaxation data on proteins. This method introduces the frequency-dependent order parameter, S(2)(omega), in order to estimate contributions to the generalized order parameter S(2) from different motional frequencies occurring on the picosecond to nanosecond time scales. S(2)(omega) is defined as the sum of a specified set of weighting coefficients from the Lorentzian expansion of the spectral density function.

Heat capacities and a snapshot of the energy landscape in protein GB1 from the pre-denaturation temperature dependence of backbone NH nanosecond fluctuations.

Protein stability is usually characterized calorimetrically by a melting temperature and related thermodynamic parameters. Despite its importance, the microscopic origin of the melting transition and the relationship between thermodynamic stability and dynamics remains a mystery. Here, NMR relaxation parameters were acquired for backbone 15NH groups of the 56 residue immunoglobulin-binding domain of streptococcal protein G over a pre-denaturation temperature range of 5-50 degrees C.

Angular variances for internal bond rotations of side chains in GXG-based tripeptides derived from (13)C-NMR relaxation measurements: Implications to protein folding.

The study of backbone and side-chain internal motions in proteins and peptides is crucial to having a better understanding of protein/peptide "structure" and to characterizing unfolded and partially folded states of proteins and peptides. To achieve this, however, requires establishing a baseline for internal motions and motional restrictions for all residues in the fully, solvent-exposed "unfolded state." GXG-based tripeptides are the simpliest peptides where residue X is fully solvent exposed in the context of an actual peptide.