Physical binding

Other immobilization methods are based on chemical and physical binding to soHd supports, eg, polysaccharides, polymers, glass, and other chemically and physically stable materials, which are usually modified with functional groups such as amine, carboxy, epoxy, phenyl, or alkane to enable covalent coupling to amino acid side chains on the enzyme surface. These supports may be macroporous, with pore diameters in the range 30—300 nm, to facihtate accommodation of enzyme within a support particle. Ionic and nonionic adsorption to macroporous supports is a gentle, simple, and often efficient method. Use of powdered enzyme, or enzyme precipitated on inert supports, may be adequate for use in nonaqueous media. Entrapment in polysaccharide/polymer gels is used for both cells and isolated enzymes. 

Substrate inhibition physical binding of substrate adsorption of substrate on resin... 

Physical binding studies (8,9,27) suggest that physical complex formation with DNA by intercalation appears to be sequence-specific. Thus, BaPT and pyrene intercalate much more strongly in poly(dA-... 

In vitro studies of DNA interactions with the reactive ben-zo[a]pyrene epoxide BPDE indicate that physical binding of BPDE occurs rapidly on a millisecond time scale forming a complex that then reacts much more slowly on a time scale of minutes (17). Several reactive events follow formation of the physical complex. The most favorable reaction is the DNA catalyzed hydrolysis of BPDE to the tetrol, BPT (3,5,6,8,17). At 25°C and pH=7.0, the hydrolysis of BPDE to BPT in DNA is as much as 80 times faster than hydrolysis without DNA (8). Other reactions which follow formation of physical complexes include those involving the nucleotide bases and possibly the phosphodiester backbone. These can lead to DNA strand scission (9 34, 54-56) and to the formation of stable BPDE-DNA adducts. Adduct formation occurs at the exocyclic amino groups on the nucleotide bases and at other sites (1,2,9,17,20, 28,33,34,57,58). The pathway which leads to hydrocarbon adducts covalently bound to the 2-amino group of guanine has been the most widely studied. 

On the basis of PAH physical binding studies (18) it has been previously suggested that BPDE adduct conformations are strongly dependent upon the DNA environment and that this may be playing a role in the varying results reported by different laboratories. A mechanism for reaction has been proposed (2) which involves initial intercalation of BPDE followed by reaction which can lead ultimately to nonintercalated complexes. This mechanism is supported by recent kinetic flow dichroism studies (69). 

Two specific suggestions concerning the role that the reversible physical binding of proximate and ultimate carcinogens derived from BP play in carcinogenesis have been made. The first is based on recognition that DNA-BPDE complex formation precedes re-... 

In order to gain more detailed information about the physical binding of hydrocarbon metabolites to DNA, studies have also been carried out with model compounds which have many of the steric and electronic properties of carcinogenic epoxides but no reactive epoxide group. The use of nonreactive model compounds permits the clear separation of physical binding interactions from reactive interactions. Benzo[a]pyrene metabolite model compounds which have been examined include 7-hydroxy-7,8,9,10-tetrahydro-BP (4), and cis (4 ) and trans-7,8-dihydroxy-7,8,9,10-tetrahydro-BP (9). 

The major goals of recent studies of the physical binding to DNA of BP and DMBA metabolites and metabolite models are to determine (1) the magnitudes of the binding constants, (2) the conformations of physical complexes which are formed and the nature of DNA binding sites, (3) how DNA structure and environment influence physical binding, (4) how the structure of hydrocarbon metabolites influences physical binding properties, (5) whether the... 

Most studies of the physical binding of hydrocarbon metabolites and metabolite model compounds have measured the effect of DNA binding on hydrocarbon fluorescence intensities, fluorescence lifetimes and UV absorption spectra Radioactive labelling has also been used, but less frequently. Spectroscopic methods are particularly convenient. These methods, especially fluorescence methods, are also very sensitive. All of the hydrocarbons in Figure 1 except the epoxides have high fluorescence quantum yields, which permit routine detection in the 10 -10 7 M concentration range. 

Intercalation of BPDE. Several groups have studied the reversible intercalative binding of BPDE to DNA. The fluorescence quantum yield of BPDE is much lower than that of BP derivatives which do not contain an epoxide group and fluorescence techniques have not been widely used to study BPDE physical binding to DNA (4). Association constants for the DNA intercalation of BPDE have been obtained by measuring red shifts in the UV absorption spectra of BPDE which occur upon the formation of intercalated complexes and from fluorescence studies (8) of the kinetics of DNA catalyzed hydrolysis of BPDE. The hydrolysis reaction is conveniently monitored by following the fluorescence of the hydrolysis product, BPT, which has a quantum yield many times greater than BPDE. 

Base Specificity of Physical Binding. To determine whether the physical binding of hydrocarbon metabolites to DNA exhibits base specificity, the binding of trans-7,8-dihydroxy-7,8,9,10-tetrahy-dro-BP was examined using fluorescence and absorption techniques (9). A comparison was also made of the varying degrees to which different synthetic polynucleotides are able to solubilize BPT... 

The Influence of DNA Structure and Environment on the Intercalation of Hydrocarbon Metabolites and Metabolite Model Compounds. The physical binding of hydrocarbon metabolites to DNA is very sensitive to DNA structure and environment. This is demonstrated by the data in Figures 4 and 5, which show how heat denaturation of DNA inhibits hydrocarbon quenching. These results are consistent with early studies which indicate that the ability of native DNA to solubilize pyrene and BP is much greater than that of denatured DNA (40). 

In addition to influencing hydrocarbon metabolite-DNA reactions, the physical binding properties of hydrocarbon metabolites covalently bound to DNA may also be important to carcinogenic activity. The covalent binding of ultimate carcinogens derived from BP and DMBA to DNA produces adducts with tt binding properties similar to those of naturally occurring nucleotides. These adducts... 

Adsorption Is the process by which some substances physically bind to the surface of a solid polar substance. 

This capacity to combine, just as with inorganic chemicals, derives from the existence of valency bonds, or links, that allow the combinations to take place. These bonds are not physical bindings like cords. They are more like magnetic attractions, although they are not actually magnetic. Each unit of carbon, like any other elemental chemical unit, is an atom, and each carbon atom has four valency bonds that can be represented as follows, although they don t actually occur like this physically ... 

Atrazine use in ecofallow usually is supplemented with other herbicides. For example, the first herbicide application to wheat stubble often uses glyphosate and 2,4-D or dicamba, with the atrazine application postponed until later in summer to coincide with the emergence of volunteer wheat, cheat, and downy brome. Atrazine can be applied with glyphosate, but antagonism with some atrazine formulations is associated with this tank mixture (Stahlman and Phillips, 1979 Wicks and Hanson, 1995) because of physical binding of inert components in the atrazine formulation with glyphosate (Ahmadi et al., 1980). Farmers know that if rainfall does not move atrazine off the wheat residue and into the soil, control of weeds, and volunteer wheat will be unsatisfactory. 

C.M. Douglas, unpublished). Demonstration of specific physical binding of MK-0911 to a fungal target remains a technical hurdle. 

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