Functional and conformational characterization of new mutants of heart fatty acid-binding protein.


Abstract

In this study we investigated the possible involvement of several amino acids (not located in the ligand-binding centre) in fatty acid binding and conformational stability of heart fatty acid-binding protein (H-FABP). We prepared recombinant human H-FABP proteins with mutations in the hydrophobic patch (Phe(4), Trp(8) and Phe(64)), portal region (Phe(16)), hinge region (Leu(66), Gly(67)), second portal region (Glu(72)) and at the protein surface (Lys(21)) respectively. Oleic acid-binding affinity and conformational stability of human H-FABP are significantly decreased or completely lost by mutation of Trp(8) or Phe(16). NMR spectra confirmed that these residues are important for the stability of the protein fold. Substitution of Phe(4) or Phe(64) resulted in less stability, but oleic acid-binding affinity was not affected. Mutation of Lys(21) had no effect on either structural integrity or fatty acid-binding affinity. Replacement of Leu(66) or Gly(67) did not affect fatty acid binding, but protein stability was reduced. Finally, mutation of Glu(72) to Ser caused no change of affinity, but NMR spectra and urea-denaturation curves showed the extremely poor stability of this mutant. In conclusion, no relationship was observed between fatty acid-binding affinity and conformational stability. Study holds ProTherm entries: 12966, 12967, 12968, 12969, 12970, 12971, 12972, 12973, 12974, 12975, 12976, 12977 Extra Details: fluorescence; NMR spectroscopy; portal region; sitedirected mutagenesis

Submission Details

ID: LDzqoY684

Submitter: Connie Wang

Submission Date: April 24, 2018, 8:44 p.m.

Version: 1

Publication Details
Zimmerman AW;Rademacher M;Rüterjans H;Lücke C;Veerkamp JH,Biochem. J. (1999) Functional and conformational characterization of new mutants of heart fatty acid-binding protein. PMID:10567233
Additional Information

Structure view and single mutant data analysis

Study data

No weblogo for data of varying length.
Colors: D E R H K S T N Q A V I L M F Y W C G P
 

Data Distribution

Studies with similar sequences (approximate matches)

Correlation with other assays (exact sequence matches)


Relevant PDB Entries

Structure ID Release Date Resolution Structure Title
1G5W 2000-11-02T00:00:00+0000 0 SOLUTION STRUCTURE OF HUMAN HEART-TYPE FATTY ACID BINDING PROTEIN
1HMR 1994-01-02T00:00:00+0000 1.4 1.4 ANGSTROMS STRUCTURAL STUDIES ON HUMAN MUSCLE FATTY ACID BINDING PROTEIN: BINDING INTERACTIONS WITH THREE SATURATED AND UNSATURATED C18 FATTY ACIDS
1HMS 1994-01-02T00:00:00+0000 1.4 1.4 ANGSTROMS STRUCTURAL STUDIES ON HUMAN MUSCLE FATTY ACID BINDING PROTEIN: BINDING INTERACTIONS WITH THREE SATURATED AND UNSATURATED C18 FATTY ACIDS
1HMT 1994-01-02T00:00:00+0000 1.4 1.4 ANGSTROMS STRUCTURAL STUDIES ON HUMAN MUSCLE FATTY ACID BINDING PROTEIN: BINDING INTERACTIONS WITH THREE SATURATED AND UNSATURATED C18 FATTY ACIDS
2HMB 1992-09-11T00:00:00+0000 2.1 THREE-DIMENSIONAL STRUCTURE OF RECOMBINANT HUMAN MUSCLE FATTY ACID-BINDING PROTEIN
3RSW 2011-05-02T00:00:00+0000 2.6 Crystal Structure of Heart Fatty Acid Binding Protein (FABP3)
3WBG 2013-05-16T00:00:00+0000 2.15 Structure of the human heart fatty acid-binding protein in complex with 1-anilinonaphtalene-8-sulphonic acid
3WVM 2014-05-25T00:00:00+0000 0.88 The 0.88 angstrom X-ray structure of the human heart fatty acid-binding protein complexed with stearic acid
3WXQ 2014-08-04T00:00:00+0000 1.6 Serial femtosecond X-ray structure of human fatty acid-binding protein type-3 (FABP3) in complex with stearic acid (C18:0) determined using X-ray free-electron laser at SACLA
4TJZ 2014-05-25T00:00:00+0000 0.87 The 0.87 angstrom X-ray structure of the human heart fatty acid-binding protein complexed with capric acid

Relevant UniProtKB Entries

Percent Identity Matching Chains Protein Accession Entry Name
90.1 Fatty acid-binding protein, heart P07483 FABPH_RAT
91.0 Fatty acid-binding protein, heart Q99P61 FABPH_ICTTR
90.2 Fatty acid-binding protein, heart O02772 FABPH_PIG
100.0 Fatty acid-binding protein, heart P05413 FABPH_HUMAN