Interaction between poly(L‐lysine⁴⁸, L‐histidine⁵²) and DNA

Abstract

SynopsisP o l y ( L y~~~, His52), a random copolypeptide of L-lysine (48%) and L-histidine (52%), was used as a model protein for investigating the effects of protonation on the imidazole group of histidines on protein binding to DNA. The complexes formed between p01y(Lys~~, His5? and DNA were examined using absorbance, circular dichroism (CD), and thermal denaturation. Although increasing pH reduces the charges on histidine side chains in the model protein, the protein still binds the DNA with approximately one positive charge per negative charge in protein-bound regions. Nevertheless, CD and melting properties of poly(Lys4*, His5*)-DNA complexes still depend upon the solution pH which determines the protonation state of imidazole group of histidine side chains. At pH 7.0, the complexes show two characteristic melting bands with a t , (46-51OC) for free base pairs and a t', (94OC) for proteinbound base pairs. The t k of the complexes is reduced to 90°C a t pH 9.2, although a t this pH there is still one lysine per phosphate in protein-bound regions. Presumably, the presence of deprotonated histidine residues destabilizes the native structure of protein-bound DNA. The binding of this model protein to DNA causes a red shift of the crossover point and both a red shift and a reduction of the positive CD band of DNA near 275 nm. This phenomenon is similar to that caused by polylysine binding. These effects, however, are greatly diminished when histidine side chains in the model protein are deprotonated. The structure of already formed poly(Lys4*, H~S~~) -D N A complexes can be perturbed by changing the solution pH. However, the results suggest a readjustment of the complex to accommodate charge interactions rather than a full dissociation of the complex followed by reassociation between the model protein and DNA.

Differential scanning calorimetric studies of the thermal stability of plasmid DNA complexed with cationic lipids and polymers

“…Previous thermal unfolding studies 2,20,21 of LIN/OC DNA have found PARG to be less effective than PLL. Factors other than ionic interactions were claimed to in¯uence the thermal stability of the complexes, 2,17,22 including the level of hydration in contact regions 18 and the ability of the polymer to bind in either the major or minor groove of DNA. 2 Several complexes were not suitable for DSC measurement because they visibly aggregated from solution.…”

Structure/Function Analysis of Peptoid/Lipitoid:DNA Complexes

“…The nearly complete exclusion of ethidium from the (A) 36 :DNA complexes indicates that this polymer binds with the tightest affinity to DNA. These results are quite different Values represent the mean of two samples of each polymer:DNA complex, except for the (APPP) 9 , (AP) 18 complexes and both lipitoid complexes, which were prepared singly. than those reported for the ability of these polymers to protect DNA from DNase I digestion, where (APP) 12 provided protection but (A) 36 did not.…”

“…To investigate the colloidal properties of the complexes formed from peptoids and lipitoids with DNA, the mean cumulant diameters of the complexes were measured using DLS ( Figure 3) and their z-potentials were obtained using PALS (Table 1). Overall, the (AII) 12 and (APP) 12 complexes possessed the smallest particle size while the (AI) 18 and (A) 36 peptoids formed the largest DNA complexes. This finding suggests that the ability of these polymers to condense DNA into small particles is mediated by factors other than the charge density of the peptoid.…”

“…Complexes were formed at a DNA concentration of 50 mg/mL (1.5 mL total volume) at charge ratios between 0.25 to 1.0 and 2.0 to 3.0 (þ/À) in purified water and were equilibrated for 10 min before measurement. Samples were prepared in duplicate except for (APPP) 9 and (AP) 18 and both lipitoids, which were prepared once for each charge ratio due to the limited amounts of these samples available. Spectra were obtained from 200 to 350 nm with a Jasco J-720 spectropolarimeter at 258C using a scan rate of 50 nm/min with a 0.1 cm pathlength cell.…”

“…The relative binding strength of peptoids and lipitoids to DNA was estimated from the ability of increasing amounts of cationic polymer to exclude the intercalation of the fluorescent probe ethidium bromide (EtBr) from DNA. Complexes containing (APP) 12 , (AII) 12 , (AP) 18 , or (AI) 18 at charge ratios less than 2:1 were least able to exclude EtBr (Figure 4). These differences were more apparent when the cationic polymer is in molar excess.…”