ABSTRACT
The morphology of tRNA was studied upon conjugation with testosterone and its aliphatic and aromatic dimers, using multiple spectroscopic methods, transmission electron microscopy (TEM) and molecular modeling. Structural analysis showed that testosterone binds tRNA through A62, A64, C60, C61, C63, G51, U50 and U59 bases. The binding affinity was testosterone dimer-aromatic>testosterone dimer-aliphatic>testosterone. The steroid loading efficacy was 35-45%. Transmission electron microscopy showed major changes in tRNA morphology upon testosterone interaction with an increase in the diameter of the tRNA aggregate, indicating encapsulation of testosterone by tRNA.
Subject(s)
Molecular Docking Simulation/methods , RNA, Transfer/chemistry , RNA, Transfer/metabolism , Testosterone/chemistry , Testosterone/metabolism , Binding Sites/physiology , Models, Molecular , Protein Structure, Tertiary , Testosterone/analogs & derivativesABSTRACT
Conjugation of DNA with testosterone and it aliphatic dimer (alip) and aromatic dimer (arom) was investigated in aqueous solution at pH 7.4. Multiple spectroscopic methods, transmission electron microscopy (TEM) and molecular modeling were used to characterize steroid-DNA binding and DNA morphology. Spectroscopic analysis showed that testosterone binds DNA via A7, A16, A17, T8, T15 and T18 nucleobases with overall binding constants Ktest-DNA=1.8 (±0.4)×104M-1, Ktest-dimeralip-DNA=5.7 (±0.7)×104M-1 and Ktest-dimer-arom-DNA=7.3 (±0.9)×104M-1. The binding affinity increases in this order: testosterone dimer-aromatic>testosterone dimer-aliphatic>testosterone. The steroid loading efficacy was 40-50%. Transmission electron microscopy showed major changes in DNA morphology as testosterone-DNA interaction occurred with increase in the diameter of the DNA aggregate, indicating encapsulation of testosterone by DNA. Modeling showed the presence of several nucleobases attached to testosterone with the free binding energy of -4.93Kcal/mol.
Subject(s)
DNA/chemistry , Dimerization , Testosterone/chemistry , Testosterone/pharmacology , DNA/metabolism , Molecular Docking Simulation , Nucleic Acid Conformation , Structure-Activity Relationship , Testosterone/metabolismABSTRACT
Due to the poor solubility of steroids in aqueous solution, delivery of these biomaterials is of major biomedical importance. We have reviewed the conjugation of testosterone and it aliphatic dimer and aromatic dimer with several carrier proteins, human serum albumin (HSA), bovine serum albumin (BSA) and milk beta-lactoglobulin (b-LG) in aqueous solution at physiological pH. The results of multiple spectroscopic methods, transmission electron microscopy (TEM) and molecular modeling were compared here. Steroid-protein bindings are via hydrophilic and H-bonding contacts. HSA forms more stable conjugate than BSA and b-LG. The stability of steroid-protein conjugates is testosterone>dimer-aromatic>dimer-aliphatic. Encapsulation of steroids by protein is shown by TEM images. Modeling showed the presence of H-bonding, which stabilized testosterone-protein complexes with the free binding energy of -12.95 for HSA and -11.55 for BSA and -8.92kcal/mol for b-LG conjugates. Steroid conjugation induced major perturbations of serum protein conformations. Serum proteins can transport steroids to the target molecules.
Subject(s)
Carrier Proteins/metabolism , Steroids/metabolism , Animals , Carrier Proteins/chemistry , Humans , Hydrogen Bonding , Hydrophobic and Hydrophilic Interactions , Lactoglobulins/chemistry , Lactoglobulins/metabolism , Molecular Docking Simulation , Protein Binding , Serum Albumin/chemistry , Serum Albumin/metabolism , Serum Albumin, Bovine/chemistry , Serum Albumin, Bovine/metabolism , Spectroscopy, Fourier Transform Infrared , Steroids/chemistryABSTRACT
A substantial part of steroids is bound to serum proteins in vivo. We report the association of testosterone and it aliphatic dimer (alip) and aromatic dimer (arom) with human serum albumin (HSA) and bovine serum albumin (BSA) in aqueous solution at physiological pH. Multiple spectroscopic methods, transmission electron microscopy (TEM) and molecular modeling were used to characterize steroid-protein binding and protein aggregation process. Spectroscopic analysis showed that steroids bind protein via hydrophobic, hydrophilic and H-bonding interactions. HSA forms more stable complexes than BSA. The binding affinity of steroid-protein adducts is testosterone>dimer-aromatic>dimer-aliphatic. Transmission electron microscopy showed major changes in protein morphology as steroid-protein complexation occurred with increase in the diameter of the protein aggregate indicating encapsulation of steroids by serum proteins. Modeling showed the presence of H-bonding stabilized testosterone-protein complexes with the free binding energy of -12.95 for HSA and -11.55 kcal/mol for BSA, indicating that the interaction process is spontaneous at room temperature. Steroid complexation induced more perturbations of BSA conformation than HSA.
Subject(s)
Serum Albumin, Bovine/metabolism , Serum Albumin/metabolism , Testosterone/metabolism , Animals , Binding Sites , Cattle , Dimerization , Humans , Hydrogen Bonding , Hydrophobic and Hydrophilic Interactions , Microscopy, Electron, Transmission , Molecular Docking Simulation , Protein Binding , Protein Structure, Tertiary , Serum Albumin/chemistry , Serum Albumin, Bovine/chemistry , Spectrometry, Fluorescence , Spectroscopy, Fourier Transform Infrared , Testosterone/chemistry , ThermodynamicsABSTRACT
The encapsulation of testosterone and it aliphatic dimer (alip) and aromatic dimer (arom) with milk ß-lactoglobulin (ß-LG) was studied in aqueous solution at pH 7.4. Multiple spectroscopic methods, transmission electron microscopy (TEM) and molecular modeling were used to characterize testosterone-ß-LG binding and protein aggregation process. Spectroscopic analysis showed that steroids bind ß-LG via hydrophobic and H-bonding interactions with overall binding constants K test-ß-LG = 5.6 (± 0.6) × 10(4)M(-1), K test-dimeralip-ß-LG = 4.8 (± 0.5) × 10(3)M(-1) and K test-dimer-arom-ß-LG = 2.9 (± 0.4) × 10(4)M(-1). The binding affinity was testosterone > testosterone dimer-aromatic > testosterone dimer-aliphatic. Transmission electron microscopy showed major changes in protein morphology as testosterone-protein complexation occurred with increase in the diameter of the protein aggregate indicating encapsulation of steroids by ß-LG. Modeling showed the presence of H-bonding stabilized testosterone-ß-LG complexes with the free binding energy of -9.82 Kcal/mol indicating that the interaction process is spontaneous at room temperature.
