Proteins are dynamic systems. Recent evidence demonstrates that they exist in a large number of conformational substates and can continuously move from one substate to another; motion of a small ligand inside a protein may be possible only through these conformational fluctuations. To test this idea, we study with flash photolysis the binding of CO to protoheme and O2 and CO to myoglobin in many different solvents. The standard evaluation of such experiments yields information only about the protein-solvent system. A novel approach is presented which permits conclusions concerning the protein: Data from all solvents are considered together, and the rates for transitions of the ligand over various barriers are studied as a function of temperature for fixed solvent viscosities. Results show that over a wide range in viscosity the transition rates in heme-CO are inversely proportional to the solvent viscosity and can consequently be described by the Kramers equation. The rates of O2 and CO in myoglobin also depend on the solvent viscosity and are most sensitive to the solvent at the lowest viscosity. Viscosity influences protein reactions even in aqueous solutions. The data dan be interpreted by a dynamic model in which transitions into and inside myoglobin are governed by fluctuations between conformational substates corresponding to closed and open pathways. Ligand motion thus is mainly controlled by gates and not by static potential barriers. Some characteristic parameters for the substates are determined, and they agree approximately with similar parameters found in Mössbauer experiments. As expected, the barrier parameters evaluated in the novel approach deviate markedly from the ones obtained by the conventional procedure. Comparison with model calculations or basic theories will be meaningful only with the new evaluation, and the method may be essential for many or possibly all biochemical reactions.

We have studied the infrared spectra of the bound and photodissociated states ofMb-"2CO and Mb-'3CO from 5.2 to 300 K. The absorbance peaks seen between 1800 and 2200 cm-1 correspond to CO stretching vibrations. In the bound state of Mb-'2CO, the known lines A0 at 1969, A1 at 1945, and A2 at 1927 cm-1, have center frequencies, widths, and absorbances that are independent of temperature between 5.2 and 160 K. Above 160 K, A2 gradually shifts to 1933 cm-'. The low-temperature photodissociated state (Mb*) shows three lines (BO, B1, B2) at 2144, 2131, and 2119 cm-' for "2CO. The absorbances of the three lines depend on temperature. Bo is tentatively assigned to free CO in the heme pocket and B1 and B2, to CO weakly bound to the heme or heme pocket wall. The data are consistent with a model in which photodissociation of MbCO leads to B1 and B2. B2 decays thermally to B1 above 13 K; rebinding to A occurs from B1. The barriers between B2 and B1 and between B1 and A are described by activation enthalpy spectra. Heme and the central metal atom in state Mb* have near-infrared, EPR, and Mossbauer spectra that differ slightly from those of deoxyMb. The observation of essentially free CO in state B implies that the difference between Mb* and deoxyMb is not due to an interaction of the flashed-off ligand with the protein but is caused by an incomplete relaxation of the protein structure at low temperatures.The reversible binding of CO to the storage protein Mb can be studied with flash photolysis (1). Experiments in which the Soret line was monitored demonstrate that the binding process involves a number ofsteps (2, 3). Here we show that monitoring the CO stretching vibration reveals additional features of the protein's interior.The active center of Mb, the heme group, is embedded in the protein (Fig. 1) (4) and the ligand binds at the central heme iron. In flash photolysis, the bound-state MbCO is photodissociated. Below 200 K the CO cannot leave the heme pocket and rebinds from there. Two states are involved in low-temperature recombination: state A, in which the CO is bound, the heme is nearly planar, and the iron atom has spin 0; and state B, in which the CO is photodissociated from the heme iron and remains in the protein pocket and the iron has spin 2. At low temperatures, the rebinding process B to A is not exponential in time. We have explained this observation by postulating the existence of conformational substates (2, 5). At low temperatures each Mb molecule is frozen into a particular substate with a specific barrier height for rebinding. From 180 to 80 K the transition occurs by an over-the-barrier Arrhenius process; below 60 K, quantum mechanical tunneling dominates (6, 7). State B (Mb*) has been studied in MbCO and CoMbCO by near-infrared (8, 9), EPR (10, 11), and Mbssbauer (12) Pentex (Kankakee, IL) was dissolved in 70% (vol/vol) glycerol in water buffered to pH 7 with 0.1 M phosphate. The sample was stirred under a CO atmosphere for several hours, reduced with sodium dithionite, and stirred for s...

A large percentage of farmers do not respond to mail surveys. To gain insight into why farmers do not respond and how to improve response rates, a three-step research design was developed. First, an initial survey, based on in-person interviews with 15 farmers, was sent to 100 farmers. Second, farmers who did not respond to this mail survey were contacted by phone to investigate the reasons for not responding. Third, based on the information from these nonrespondents, the survey instrument was revised and sent to 3,990 U.S. farmers. Our studies show that the period in which the survey is sent is a crucial factor in the willingness to participate, along with the form and amount of compensation, the sender of the questionnaire, and the perceived length of the questionnaire.

Binding of carbon monoxide to the separated alpha and beta chains of hemoglobin, with and without bound p-mercuribenzoate, has been measured at temperatures from 5 to 340 K for times 2 mus to 1 ks using flash photolysis. All four proteins exhibit three different rebinding processes. The data are interpreted by a model in which the carbon monoxide, moving from the solvent to the binding site at the ferrous heme iron, encounters three barriers. The temperature dependences of the three processes yield activation enthalpies and entropies for the three barriers for all four proteins. Binding at temperatures below about 200 K is nonexponential, implying that the innermost barrier has a distribution of activation enthalpies. The distributions for the four proteins have been determined. At temperatures below 30 K, the CO binding rates approach finite low-temperature limits; binding thus proceeds by quantum-mechanical tunneling. Invoking a simple model, the widths of the innermost barriers are extracted from the measured tunneling rates. The experimental parameters are correlated with structural features of the hemoglobin chains and compared with previously published data on myoglobin and protoheme. A correlation is established between the height of the innermost barrier and the equilibrium CO pressure.

We investigate the influence of weather anomalies on net migration in the Eastern United States using a county-level panel for the period from 1970 to 2009. One major mechanism is through the effect of weather on agricultural yields, which we examine in further detail using an instrumental variables approach. Our preferred model uses the seasonality of the sensitivity of corn yields to extreme heat over the growing season, which peaks during corn flowering, as instrument. The reduced-form estimate of the migration response to extreme heat closely mirrors the seasonality of corn yield. Our IV approach will provide an unbiased estimate of the responsiveness of outmigration to yield unless other determinants of migration, such as peoples direct preference for weather, perfectly align with the pattern of corn flowering. This is unlikely given that the exact dates of corn flowering vary from year to year. Our estimated semi-elasticity ranges from-0.3 to-0.4 depending on the chosen time trend, i.e., a one percent change in yields leads to an opposite 0.3-0.4 percentage point change in the net migration rate. The migration response is strongest for young adults and not significant for senior citizens. Extrapolating from this relationship, we project that climate change would induce significant outmigration in the U.S. Corn Belt.