Surfactant-triggered disassembly of electrostatic complexes probed at optical and quartz crystal microbalance length scales.

A critical advantage of electrostatic assemblies over covalent and crystalline bound materials is that associated structures can be disassembled into their original constituents. Nanoscale devices designed for the controlled release of functional molecules already exploit this property. To bring some insight into the mechanisms of disassembly and release, we study the disruption of molecular electrostatics-based interactions via competitive binding with ionic surfactants. To this aim, free-standing micrometer-size wires were synthesized using oppositely charged poly(diallyldimethylammonium chloride) and poly(acrylic acid) coated iron oxide nanoparticles. The disassembly is induced by the addition of sodium dodecyl sulfates that complex preferentially the positive polymers. The process is investigated at two different length scales: the length scale of the particles (10 nm) through the quartz crystal microbalance technique and that of the wires (>1 µm) via optical microscopy. Upon surfactant addition, the disassembly is initiated at the surface of the wires by the release of nanoparticles and by the swelling of the structure. In a second step, erosion involving larger pieces takes over and culminates in the complete dissolution of the wires, confirming the hypothesis of a surface-type swelling and erosion process.

Separation and evaluation of the covalent and noncovalent interactions which contribute to the binding of pyridoxal 5'-phosphate to D-serine apodehydratase.

The equilibrium constant (KX) for the reaction D-serine dehydratase + pyridoxamine-P in equilibrium KX D-serine apodehydratase: pyridoxamine-P + pyridoxal-P was determined. At 25 degreees, pH 7.80, KX increases from 5.4 times 10-minus 5 to 21 times 10-minus 5 as T/2 is increased from 0.33 to 0.66. A value of 1.3 times 10-minus 4 M at 25 degrees, pH 7.80, T/2 0.33 for the equilibrium constant (KPMP) for dissociation of pyridoxamine-P from D-serine apodehydratase was determined from the ratio of the equilibrium constant for dissociation of pyridoxal-P from holoenzyme to KX. Pyridoxamine-P and the thiazolidine, formed from pyridoxal-P and cysteine, were found to have similar affinities for D-serine apodehydratase. Using the affinities of these derivatives as a measure of the noncovalent interactions between cofactor and protein, it was possible to estimate the contribution of the Schiff base linkage to the stability of the complex formed between pyridoxal-P and protein. The covalent Schiff base linkage in the holoenzyme was found to be no more stable than the Schiff base linkage formed between 6-aminocaproic acid and pyridoxal-P. The contribution of noncovalent interactions to the stability of the cofactor-protein complex was shown to be at least 20 to 40 times greater than the contribution of the covalent Schiff base linkage.

Resolution of D-serine dehydratase by cysteine. An analytical treatment.

A general method is presented for analysis of the resolution of pyridoxal-P-requiring enzymes by carbonyl reagents. The method is useful for accurately determining the very small equilibrium constants (KP) which characterize the dissociation of cofactor from many pyridoxal-P-requiring enzymes. The analysis also establishes the minimum number and relative stabilities of distinct enzymic species involved in the resolution process. Analysis of the resolution of D-serine dehydratase by L-and D-cysteine resulted in the establishment of an enzyme bound thiazolidine derivative as an intermediate in the pathway for resolution. The over-all equilibrium constant (KR) for the reaction, D-serine dehydratase + cystein in equilibrium KR thiazolidine derivative +D-serine apodehydratase was determined. At pH 7.80, T/2 0.33, 25 degrees, KR equal to 1.08 times 10-minus 3. A value of 7.0 nM for the equilibrium constant for the dissociation of D-serine dehydratase to apoenzyme and free pyridoxal-P was determined from the ratio KR/KT, where KT is the equilibrium constant for the formation of a thiazolidine derivative from free pyridoxal-P and cysteine. An estimate of 14 nM for KP was also obtained from partial resolution of D-serine dehydratase by high dilution. The difficulties associated with this direct determination of KP from the dependence on the enzyme concentration of the activity of very dilute solutions of enzyme are discussed.

Reactions of pyrzdoxal 5'-phosphate, 6-aminocaproic acid, cysteine, and penicilamine. Models for reactions of Schiff base linkages in pyridoxal 5'-phosphate-requiring enymes.

Schiff base formation, transaldimination, and reaction with aminothiols are important reactions which occur on the surface of pyridoxal-P-requiring enzymes. As a first step in assessing the role of the protein in these reactions, models for these reactions were studied in the absence of enzyme at 25 degrees, gamma/2 0.28. The reaction of 6-aminocaproic acid with pyridoxal-P to form the Schiff base N6-(P-pyridoxylidene)-aminocaproic acid was studied as a model for formation of Schiff base on the holoenzyme...