HYDRATION QUANTIZATION OF RECEPTOR BINDING SITES














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Author
Introduction
Water
Protein Structure
Carboxypeptidase A
Alpha Chymotrypsin
Insulin
Ribonuclease A
Receptor Sites
Double-Helix B-DNA
References


INTRODUCTION

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INTRODUCTION

Historically, each water molecule in the liquid state has been considered to be dynamically hydrogen-bonded in tetrahedral configurations to three or four other water molecules.7 Bonds are weak (1.3 to 2.8 kcal/mole) and last only about 10-11 seconds but continually tie the molecules together in clusters.8 The extremely high melting and boiling points of water are considered due to this type of hydrogen bonding.7 However, recent high-speed neutron irradiation studies have provided evidence that, at any instant, water molecules in the liquid state hydrogen-bond more-strongly together to form trimers with two water molecules hydrogen-bonded to a central molecule.9  Molecular orbital calculations back in the 60’s and 70’s forecast that a trimer, with a hydrogen-bond length of about 2.76 Angstroms, would be the largest stable unit in liquid water10 and, in 1972, the trimer and a linear tetramer were identified by X-ray scattering on the surface of liquid water.11 However, the nature of hydrogen bonding in the trimer and longer ordered elements of hydration which form on hydrophobic surfaces remained a mystery until 1999 when Dr. Isaacs at Bell Labs concluded, based on X-ray analysis, that hydrogen bonding in ice is “covalent ” with a strength of 4.7 to 8.2 kcal/mole.12 As mentioned above, additional evidence for two different types bonding between water molecules was provided recently when Professor Stanley, in a detailed mathematical analysis of the properties of liquid water, concluded that water, in the liquid state and on surfaces, is composed of two different density-forms of water.6

 Tetrahedral hydrogen bonding, which is the primary form present within and on thermodynamically-stable forms of natural molecules (including proteins), involves the columbic attachment of water molecules to each other and to other polar molecules at charge-points on their surfaces.7 Distances and angles of bonding vary considerably to provide for thermodynamic stabilization.7  Covalent hydrogen-bonding, on the other hand, involves the quantized overlap of electron-clouds of adjacent water molecules around a central proton - similar to the covalent bonding between carbon atoms.12 Covalent hydrogen bonds are stable in ice but the trimer and longer linear elements which form on hydrophobic and ionic surfaces in liquid water are unstable and last only about 10-12 seconds.13

 X-ray diffraction from the surface of liquid water at 25oC produces weak peaks for the trimer at 4.5A and a linear tetramer at 6.8A (2.76 Angstroms between oxygens), with a major peak at 2.9 Angstroms for most of the molecules.11 At 2.9 Angstroms, most water molecules are far enough apart to permit rotation but close enough for charges on their surfaces to draw them into coordinated point-charge hydrogen-bonding and intermittent covalent bonding. Although trimers and tetramers represent only a small fraction of surface water, by forming rapidly and repetitively, they may contribute to the high surface tension. 

 If oil is on the surface of water, surface tension increases even more and the oil molecules lose freedom. Instead of bending, twisting and turning in the liquid state, oil molecules adjacent to water lose entropy and energy and are forced to align parallel to each other in layers.4,16 At the same time, molecular orbital calculations indicate that water molecules at the interface assemble into short covalent linear and cyclic forms parallel to the surface and in preferred orientations.17 As a water molecule moves from the dynamic liquid state to the surface state and covalently-bonds more strongly with two neighbors, it loses 8 to 10 kcal/mole of energy to adjacent water molecules but, as it moves rapidly (by a rotation)14 back into the liquid state, it absorbs similar quantized units of energy from the surface and drives molecules within that surface toward lower energy and greater order.4,15

 Thus, it is the spontaneous and rapid movement of water molecules from ordered covalently-bonded states on hydrophobic and ionically-ordered surfaces to more mobile, higher-energy point-charge states which drives more massive, slower-moving molecules, like those in gasoline and oil, toward lower energy and higher order.14,15,16 In fact, recent studies indicate that movements and energy changes in water molecules adjacent to large ions and ordering surfaces are based on Quantum Mechanics, not Newtonian Physics – they “jump” from one energy-state to the next.18  Back in 1944, it was Erwin Schrodinger, one of the founders of quantum mechanics, who concluded in his little book, What is Life?, that it is this unidirectional movement of energy from natural molecules into water which drove early random molecular systems toward order and life - the opposite direction from what one would expect based on the Second Law of Thermodynamics.19

 Of course, it is this same unidirectional transfer of quantized units of energy from hydrocarbon surfaces of polypeptides to water, as they are released from ribosomes, which not only drives folding and assembly but, by repetitively forming in particular orientations on surfaces, may assist in directing assembly and function.20  During the earliest phases of molecular evolution, when polypeptides most likely were produced at random, those which could fold spontaneously into thermodynamically-stable forms and release unstable covalently-ordered water, survived - those which could not fold into stable functional forms were chewed up by lytic enzymes and ribozymes. 

 However, trimer and linear-element formation may be responsible for another critical property of water. As pure liquid water cools, mean bonding distances between the molecules decrease and density increases until the temperature reaches 4oC; then[J1] , as the temperature is lowered to 0oC, the molecules appear to move away from each other and the density decreases.7 Often, this density decrease has been attributed to the formation of pseudo-forms of ice.6,7  However, water, in contact with a surface where atoms are in hexagonal patterns, like those in ice and iodine crystals, crystalizes immediately at 0oC. Since pure liquid water in a clean glass container can be cooled to as much as 30 degrees below zero C without crystallizing,21 the two-dimensional hexagonal forms in ice cannot be present below 4oC. Instead, trimers, which extend the hydrogen-bonding of water molecules around them, most likely form with increasing frequency and longer half-lives. As liquid water is super-cooled below 0oC, viscosity increases as the frequency of trimer formation and lengths of linear elements increase until, at -40oC, crystallization occurs immediately to yield a form of ice called “cubic” 21 in which all of the molecules are in linear elements and covalently-linked at a distance of 2.75 Angstroms.22

 Cubic ice is called the “kinetic product” because it forms most rapidly as orbitals overlap a central proton.23 However, it is unstable at atmospheric pressure and, as it warms to 0oC, it isomerizes to the thermodynamically more-stable “hexagonal” form in which the molecules are in both linear and non-linear forms at distances of 2.75A to 2.84A.22

 Although the order in water on surfaces of proteins, nucleic acids and carbohydrates is displayed by proton magnetic resonance as the doublet peaks of ice rather than the singlets of liquid water,24 molecular surfaces are so dynamic that has been impossible to visualize the precise structure of the order in water. However, ultra-high-speed 4D crystallographic analysis of water on the ridged hydrophobic surface of graphite by the late Professor Zewail and his group at CIT displayed it as linearly ordered, hexagonal layers with cubic patterning between the layers – the same as in cubic ice.25  

 Thus, as polypeptides emerge from ribosomes, the probability is high that extremely short-lived covalent linear elements of hydration form on hydrophobic surfaces and between charge centers - that it is this unstable covalent surface water which, by spontaneously moving from order toward disorder, drives those regions into more ordered, internally-bonded, coils and beta-sheets.4,15 At the same time, it is small peptides (like glycine and serine), which, by continually hydrogen-bonding dynamically with surface water, disrupt linear-element formation to permit chains the mobility to direct ordering regions into lower energy, thermodynamically more stable assemblies.2,4

 As water-soluble proteins form, most peptides with hydrocarbon side-chains are left inside forming an anhydrous core while peptides which hydrogen-bond directly with water are left on the surface disrupting coordinated hydration order to increase stability and solubility.2 However, selective regions on surfaces contain peptides which continue to induce the formation of covalent linear elements to direct proteins into functional assemblies and substrates and regulator molecules into binding sites.26 In fact, as enzyme and receptor sites open and close in response to the dynamics of surface water, linear covalent elements of hydration most likely form within those spaces.20 Half-lives of these elements are too brief to provide stability but, if substrates or regulator molecules (with similar dimensions and binding properties) are drawn into the sites to provide thermodynamic stability, proteins shift into activated conformations and perform vital functions. Based on this concept of order and disorder in surface water, it should come as no surprise that most regulator molecules, like hormones and neurotransmitters, mimic the dimensions and hydrogen-bonding properties of transient quantized linear elements of hydration.20

 Indeed, it is unfortunate that surface water, although extremely important for natural structure and function, is never displayed.  For example, the classical structure of DNA is never pictured as hydrated but the X-ray crystallographic pattern which was used by Watson and Crick to assemble their helical model, was obtained by Rosalind Franklin by spraying a crystalline sample with water.27 Only if surrounded by at least 13 “ice-like” water molecules per base-pair, does DNA adopt the helical structure that has become the logo for modern molecular biology.28 Only by transiently-forming linear elements of six to seven molecules of water between the anionic oxygens of surface phosphates across the wide groove and three to four across the narrow groove to delocalize the high negative charge, is the structure stabilized in the B Form.20  Dehydration converts DNA into the A Form which does not exist in nature.28

 For the past century, attention of the scientific community has been directed to the identification of structure for natural molecules - now it is time to reveal that it is surface water which not only provides for spontaneity and order but the quantized spatial criteria for the selection of the molecules of life.20  Although tetrahedral hydrogen bonds can vary in lengths and angles in thermodynamically-stable states, the standard hydrogen-bond length of 2.76A and angle of 109.5o (as found by molecular orbital calculations and in trimers and tetramers on the surface of liquid water at 25oC)8,9,10 are used as covalent linear elements in this article to fill spaces and stabilize molecules in transition states.

 Companion web sites, www.proteinhydration.com and www.molecularcreation.com, provide more detail on surface hydration as well as a description of the role surface water may have played in the evolution of natural molecules.

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