STRUCTURAL AND DYNAMIC CHARACTERIZATION OF THE AROMATIC-AMINO-ACIDS OF THE HUMAN-IMMUNODEFICIENCY-VIRUS TYPE-I NUCLEOCAPSID PROTEIN ZINC FINGERS AND THEIR INVOLVEMENT IN HETEROLOGOUS TRANSFER RNA(PHE) BINDING - A STEADY-STATE AND TIME-RESOLVED FLUORESCENCE STUDY

The steady-state and time-resolved fluorescence properties of two zinc -saturated 18-residue synthetic peptides with the amino acid sequence of the NH2-terminal (NCp7 13-30 F16W, where the naturally occurring Ph e was replaced by a Trp residue) and the COOH-terminal (NCp7 34-51) zi nc finger domains of human immunodeficiency virus type I nucleocapsid protein were investigated. Fluorescence intensity decay of both Trp 16 and Trp 37 residues suggested the existence of two fully solvent-expo sed ground-state classes governed by a C = 2.2 equilibrium constant. T he lifetimes of Trp 16 classes differed from those of Trp 37 essential ly because of differences in nonradiative rate constants. Arrhenius pl ots of the temperature-dependent nonradiative rate constants suggested that the fluorescence quenchers involved in both classes and in both peptides were different and the collisional rate of these quenchers wi th the indole ring was very low, probably because of the highly constr ained peptide chain conformation. The nature of the ground-state class es was discussed in relation to H-1 nuclear magnetic resonance data. U sing Trp fluorescence to monitor the interaction of both peptides with tRNA(Phe) we found that a stacking between the indole ring of both Tr p residues and the bases of tRNA(Phe) occurred. This stacking constitu ted the main driving force of the interaction and modified the tRNA(Ph e) conformation. Moreover, the binding of both fingers to tRNA(Phe) wa s noncooperative with similar site size (3 nucleotide residues/peptide ), but the affinity of the NH2-terminal finger domain (K = 1.3 (+/- 0. 2) 10(5) M-1) in low ionic strength buffer was one order of magnitude larger than the COOH-terminal one due to additional electrostatic inte ractions involving Lys 14 and/or Arg 29 residues.