4761
J. Phys. Chem. 1991, 95, 4761-4765 Optical Measurements. Absorption spectra were taken on a Varian Cary 210 spectrophotometer. Emission spectra were recorded on an Aminco SPF 500 fluorimeter working in ratio mode with an excitation and emission bandpass of 1 nm. The lifetimes were measured by the time-correlated single-photon counting technique. The instrument response function was reconstructed from the measured fluorescence decay of dimethylPOPOP. Laser characteristics and other details have been described el~ewhere.'~
Acknowledgment. This work was supported by the Belgian National Science Foundation (FNRS).We thank this foundation for a fellowship to P.V. and the IRSIA foundation for a fellowship to B.L. We are grateful to Professor F. C. De Schryver and to Dr. M. Van Der Auweraer, who allowed us to make lifetime measurements. We acknowledge Professors F. C. De Schryver, M. Van Der Auweraer, L. Eberson, G. B. Schuster, and J. Fastrez for helpful discussions.
Nitrogen-Protonation Mlcroequlllbrla and C(2)-Deprotonatlon Mlcroklnetlcs of Hlstidlne, Histamine, and Related Compounds Bila NoszQltand Dallas L. Rabenstein* Department of Chemistry, University of California, Riverside, California 92521 (Received: November 5, 1990)
The thermodynamics of nitrogen protonation and the kinetics of deprotonation of the C(2) carbon of the imidazolium ring were studied in seven imidazole derivatives. Equilibrium constants for protonation of the amino and imidazole nitrogens were determined by pH-metry and rate constants for substitution of deuterium for the C(2) hydrogen were measured by 'H NMR over the temperature range 298-333 K. The results revealed relationships between the logarithm of the product of the equilibrium constant for imidazole N protonation and the rate constant for C(2) substitution and the logarithm of the equilibrium constant for imidazole N protonation. By use of these relationships, microscopic equilibrium constants were determined for the N protonation and microscopic rate constants for the C(2) deprotonation of histamine, histidine, and histidylglycine. The interactivity parameter for amino and imidazole protonation, which could not be determined previously due to the predominance of amino over imidazole N protonation, is virtually the same for these three molecules, whereas the amino/imidazole basicity ratio significantly increases in the above order. Microscopic activation energies for substitution of deuterium for the C(2) hydrogen in these compounds were determined from the temperature dependence of the microscopic substitution rate constants.
Introduction The imidazole ring plays an important role in biological chemistry. In peptides and proteins, the imidazole ring of histidine residues is the only moiety that changes its state of protonation in the pH range of most biological fluids. This feature provides intra- and intermolecular binding versatility and conformational flexibility for histidine-containing biopolymers. Also, the imidazole nitrogens are among the strongest electron pair donors to metal ions, and their Lewis basicity depends on the protonation state of the rest of the molecule. Protonation of the imidazole group of histamine, an important molecule in immunochemical reactions, is considered to be essential for the storage of histamine in mast cell granules.'-3 The presence of the imidazole and aliphatic amino groups in histamine and histidine results in some similarities in their binding to other biomolecules and to metal ions. For histamine and histidine, it has long been recognized that the amino nitrogen is more basic than the imidazole nitrogen and consequently that the first protonation occurs predominantly a t the amino nitrogen, as represented by the upper pathway in the microscopic protonation scheme in Figure 1. Some protonation does occur first at the imidazole nitrogen; however, the amount is too small to detect by standard methods for characterizing protonation,equiiibria. Hence, despite the importance of histamine and histidine in biological chemistry, their microscopic protonation equilibria have not been characterized. Rather, their acid-base equilibria have been characterized in terms of macroconstants. Macroconstants determined at a variety of temperatures and ionic strengths have been compiled in reference booksC6 and critical The metal ion binding properties of histidine and histamine have been reviewed from the thermodynamic, structural, and kinetic points of ~iew,~JO including the nonexperimental estimation of histidine microconstants. Special attention has been 'Permanent address: Department of Inorganic and Analytical Chemistry,
L. EBtvBa University, 1518 Budapest 112,P.O. Box 32, Hungary.
0022-3654/91/2095-4761$02.50/0
paid to the imidazole N ( 1)-N(3) proton tautomerism in histidine"*'* and histamine" as well as the populati~n'~*'~ and basicity16 of histidine rotamers. Another important feature of the imidazole ring is that the carbon-bound C(2)-proton exchanges for deuterium in D20. The exchange rate has been measured for several and ~~
~
(1) Lagunoff, D. Biochemistry 1974, 13, 3982-3986. (2) Lagunoff, D.In Bronchial Asrhma: Mechanism and Therapeutics; Weiss, E. B., Segal, M. S.,Stein, M., Eds.; Little, Brown, and Co: Boston, 1985;p 236. (3) Rabenstein, D.L.; Ludowyke, R.; Lagunoff, D,Biochemistry 1987, 26, 6923-6926. (4) Sillen, L. G.; Martell, A. E. Stability Consrants of Metal-Ion Complexes, I , II; The Chemical Society: London; Special Publication No. 17and 25, 1964 and 1970. (5)Sergeant, E. P.; Dempsey, B. Ionization Consrants of Organic Acids in Aqueous Solution; Pergamon Press: Oxford, U.K., 1979. (6) Perrin, D.D.Stability Constants of Meral-Ion Complexes Part E . Organic Ligands; Pergamon Press: Oxford, U.K., 1979. (7) Martell, A. E.; Smith, R. M. Critical Stability Constants Vol. I : Amino Acids; Plenum Press: New York, 1974. (8) Smith, R. M.; Martell. A. E. Critical Srabiliry Constants Vol. 2: Amines; Plenum: New York, 1975. (9) Sundbcrg, R.J.; Martin, R. 8. Chem. Rev. 1974. 74. 471-517. (IO)Martin, R. 8. Metal Ions in Biological Systems; Siege], H . , Ed.; Marcel Dekker: New York, 1979;Vol. 9,pp 1-39. (11) Tanokura, M. Biochim. Biophys. Acta 1983, 742, 576-585. ( 1 2 ) Tanokura, M.Biopolymers 1983,22, 2563-2576. (13)Weinstein, H.;Chou, D.; Johnson, C. L.; Kang, S.;Green, J. P.Mol. Pharmacol. 1976, 12, 738-745. (14)Espersen, W. G.;Martin, R. B. J. Phys. Chem. 1976,80, 741-745. (15)Weinkam, R. J.; Jorgensen, E. C. J . Am. Chem. Soc. 1973, 95. 6084-6090. (16) Fujiwara. S.;Ishizuku, H.; Fudano, S . Chem. Lett. 1974, 1281-1284. (1 7) N d 1 . B.; Scheller-Krattiger, V.; Martin, R. B. J . Am. Chem. Soc. 1982, 104, 1078-1081. (18)Takeuchi, Y.; Kirk, K. L.; Cohen, L. A. J. Org. Chem. 1978, 43, 3570-3578. (19) Elvidge, J. A.; Jones, J. R.;Salih, R.;Shendala, M.; Taylor, S.E. J. Chem. SOC.,Perkin Trans. 2 1980, 447-451.
0 1991 American Chemical Society
4762 The Journal of Physical Chemistry, Vol. 95, No. 12, 1991
NoszAl and Rabenstein
H
D
D
XCH-CH?
k,
ND;
'"'I:>H
"$"-
N D
+OD
ND;
D
D
H
(N,lm+)
ky
D
(N+.lm+)
Figure 2. Reaction pathways of the C(2) proton-deuterium exchange in D20solutions.
H
The relationships between the macro- and microconstants are given in eqs 5 and 6.
K 1 = kN + ktm
f12 = K l K 2 = kNk@ = k"kL Figure 1. Microscopic and macroscopic protonation equilibria for the amino and imidazole groups of histidine, histamine, and histidylglycine. For histidine, X = -0OC-; for histamine, X = H-; for histidylglycine, X = -OOCCH,HNCO-.
p ~ r i n ederivatives l ~ ~ ~ ~ and utilized to determine the solvent accessibility of histidine in globular proteins. The exchange rate also depends on the protonation state of the basic groups in the molecule, and thus it can be characterized correctly only in terms of microscopic rate constants. Such values have been determined for 3-methylhistidine,17 but no data are available for histidine and histamine. In this paper, we present microscopic constants for both the N-protonation equilibria and the C(2)-proton-~ubstitutionrate of histidine, histamine, and histidylglycine. The constants were determined by a new method, which is suitable for the parallel determination of thermodynamic and kinetic parameters of imidazole derivatives. We also calculated the microscopic activation energies, which are the first parameters of this kind to be reported.
Theory The microscopic and macroscopic protonation equilibria of the common moiety of histidine, histamine, and histidylglycine are shown in Figure 1, The N and Im abbreviations stand for the
histidylglycine
amino and imidazole group, respectively. Superscript + indicates that the group in question is protonated. Accordingly, N+,Im and N,Im+ represent the two protonation isomers of the monoprotonated forms. Microconstants kN,k", k K , and @' can be expressed in terms of microspecies and hydrogen ion concentrations as follows:
[N+,lm+] krm = [N,Im+][H+]
(3) (4)
(20) Thomas, G. J.; 122-124.
Ferreira, S. A. J.
Roman Specrrosc. 1982, 12,
(6)
Microscopic acid-base equilibrium constants have been determined for several amino acids and other biomolecules by combining information from two experimental techniques.21-26 One of the experimental techniques is always pH-metry. The other spectroscopy. Spectroscopic is generally UV21-23or NMR24*25 methods can be used when (i) the minor protonation isomer OCCUA in sufficiently high concentration and (ii) the protonation processes of at least one of the two groups can be selectively monitored by the spectroscopic method. When either of these conditions is not met, chemically modified compounds have to be used as models, preferably of the minor protonation i s ~ m e r . ~ ~ - ~ ~ For histamine and histidine, the upper protonation pathway via the N+,Im protonation isomer in Figure 1 overwhelmingly predominates, and consequently the macroconstants K I and Kz are essentially equal to the microconstants kN and k k , respectively. In turn, the concentration of the N,Im+ minor protonation isomer is insufficient for the pH-metric-spectroscopic determination of the ktmand k k microconstants. The concentration ratio of the protonation isomers is independent of both the pH and the total concentration
-IN'JmI =[N,Im+]
H
X = -OOC-. histidine X = H-, histamine X = -OOC-CH&INOC,
(5)
kN k"
(7)
Since kN > ktm,under no solution conditions can the microspecies N,Im+ be the predominant species. Concerning possible model compounds for the determination of microconstants, they should bear a permanent positive charge on the imidazole ring. Even if such compounds were available [e.g., N,N'-dimethylimidazole derivatives] they would certainly lack the hydrogen-bonding characteristics of the original molecules. For these reasons, the microconstants of histidine, histamine, and related compounds cannot be determined with any of the methods previously used to measure microconstants. In this study, we have determined microconstants for histidine, histamine, and histidylglycine indirectly from the rate of substitution of the imidazole C(2) hydrogen by deuterium in D20. This exchange involves reaction with the OD- ion and takes place exclusively when the imidazole ring is protonated."-20 The two possible reaction pathways are shown in Figure 2. N+,Im+ and N,Im+ are the reactive microspecies and k++ and kw the corresponding second-order microscopic rate constants in the two reaction pathways. The overall rate equation is -dCH/dt = k++[N+,Im+][OD-] + ko+[N,Im+][OD-] (8) Benesch, R. E.; Benesch, R. J. Am. Chem. Soc. 1955,77,5877-5881. Martin, R. B. J. Phys. Chem. 1971, 75, 2657-2661. Kiss, T.; T l h , B. Talanra 1982, 29, 539-544. Rabenstein, D. L.; Sayer, T. L. Anal. Chem. 1976, 48, 1141-1146. "XU, 8.;SBndor, P. Anal. Chem. 1989,61, 2631-2637. Martin, R. B. Metal Ions in Biological Sysrems; Sigel, H . , Ed.; Marcel Dekker: N e w York, 1985; Vol. 19, p 19. (21) (22) (23) (24) (25) (26)
The Journal of Physical Chemistry, Vol. 95, No. 12, 1991 4163
Microequilibria and Microkinetics of Histidine where CHis the total concentration of all species containing hydrogen a t the C(2) position. The concentrations of N+,Im+ and N,Im+ can be expressed by means of protonation (more precisely 'deuteration") constants and deuterium ion and microspecies concentrations: -dCH/dt = k++&[N,Im] [D+I2[OD-] + ko+klm[N,Im][D+][OD-] (9) The concentration of the N,Im microspecies is equal to the product of its mole fraction (aNlm) and the total concentration (CH), whereas the product [D4][OD-] is Kw, which leads to
Integration and rearrangement of eq 10 yields
In (CO/CH)/aN,lmKwt
k++82[D+1 + kO+k"
(11)
where CO = CHat t = 0. The left-hand side of eq 1 1 can be determined by NMR and potentiometric measurements. Since only the C(2) proton exchanges for deuterium, the Co/CH ratio can be followed by monitoring the relative intensities of the resonances for the C(4) and C(2) protons as a function of time (for ring numbering see Figure 1). aN,Im can be calculated from the macroconstants and the D+ concentration: aN,lm
= 1 /(I
+ KI[D+l + 82[D+12)
The products k++B2 and ko+kImcan then be determined from measurements at two or more pH (pD) values. Since potentiometric measurements provide the macroconstant b2, the rate constant k++ can be calculated from the value obtained for k++O2. However, neither kW nor k" is known, so their individual values can not be determined without additional information. Earlier studies on a wide variety of imidazole derivativesI7J8 revealed a high correlation between the N basicity and the C(2) proton exchange rate: the lower the imidazole N basicity, the faster the C(2) proton exchange. A plot of log (rate constant) versus the corresponding log (equilibrium constant) had a correlation coefficient of nearly -1. We anticipated a similar correlation between the corresponding log (rate constant X equilibrium constant) and log (equilibrium constant) values. The existence of such a correlation was verified by studying systems where the appropriate equilibrium constants for imidazole protonation are known. In one group of systems studied, rate constant k++ could be calculated direct1 from kinetic and potentiometric data since the microconstant kN Ym is essentially equal to the macroconstant K2, as discussed above for histamine and histidine. In the second group of systems studied, the molecules contained only the imidazole group. Thus, in these cases, C(2)-hydrogen substitution is characterized by a single rate constant k+, which could be calculated by using eq 12 where K is the imidazole protonation In (Co/CH)/almKwt = k+K
(12)
constant. We studied seven imidazole derivatives, each at three elevated temperatures, and found correlation coefficients of 0.997 or larger between the log (rate constant X equilibrium constant) and log (equilibrium constant) parameters at every temperature. On this basis, the experimental ko+k" products (eq 11) for histamine, histidine, and histidylglycine were separated into kW and k". The temperature dependence of the kW and k++ microscopic rate constants provided the activation energies Eo+ and E++and that of the klm microconstants allowed extrapolation to ambient temperature. The other microconstants were then calculated by using eqs 5 and 6 and the appropriate macroconstant values. Finally, by use of the interactivity parameters (log kN - log kk = log k" - log k E = A log kN+,) the 25 OC equilibrium data were converted from D 2 0 to H 2 0 . Experimental Section Histidylglycine, N-acetylhistidine, imidazole-4-acetic acid, L-8-imidazolelactic acid, 4(5)-methylimidazole, and potassium deuteroxide were obtained from Sigma Chemical Co.; L-histidine
TABLE I: Protonation Macroconstants in D z F 298 K 313 K 323 K
histidylglycine histidine
8.09 6.39 9.70
histamine
6.60 10.63
N-acetylhistidine imidazole-4-aceticacid L-8-imidazolelactic acid 4-methylimidazole
6.72 7.46 7.71
7.67 8.10
7.75 6.03 9.30 6.24 10.18
6.43 7.14 7.33
7.55 5.83 9.12
333 K 7.32 5.66 8.19
6.12
5.90
9.88
9.55 6.10 6.68 6.95 6.98 7.33
6.31 6.90
7.16
7.34
7.23
7.75
7.52
'Values are reported in log K units. For histidylglycine, histidine, and histamine, values are reported for log K, and log K1, where log K, is the larger of the two values. Constants below log K = 5 are neglected here. b I= 2.0 mol dm-' (KCI). 'Reported log K values are the average of values obtained from 1 to 4 titrations; the estimated uncertainty is 0.01-0.04 log K units based upon the calculated standard deviations and the temperature reliability of the calibration buffers and the emf cell. was obtained from Calbiochem, histamine dihydrochloride from Reanal, D 2 0 from Icon Services Inc., and DCl from Aldrich. All chemicals were of analytical reagent grade and were used as received. The macroconstants for all seven compounds were determined by potentiometric titration of D 2 0 solutions at 298,313,323, and 333 K. The macroconstants for histidylglycine, histidine, and histamine were also determined in H20at 298 K. Concentrations of 0.15-0.2 mol dm-' were used in both the kinetic and potentiometric experiments. The ionic strength was held constant a t 2.0 mol dm-', using KCI as auxiliary electrolyte. The initial acid (DCI or HCl) and base (KOD and KOH) concentrations were 0.33 and 1.0 mol d d , respectively. The titrations were carried out using an Orion 701A pH meter, equipped with an Ingold combination microelectrode (6030 No. 3) and Mettler DV automatic buret. All pH (pD) data are pH meter readings based upon Leeds and Northrup buffer solutions of pH = 6.865 (6.838, 6.833,6.836) and pH = 9.180 (9.068,9.011,8.962) buffers (entries in brackets are buffer pH values at 313, 323, 333 K). Kinetic experiments were performed at 3 13, 323, and 333 K. Samples in NMR tubes were kept a t 313, 323, and 333 K in a water bath for 8-12, 3-5, and 1-1.8 h, respectively. IH N M R spectra were then obtained at 500 MHz with a Varian VXR 500s spectrometer, operated in the pulseFourier transform mode. A 90° pulse angle and a spectral range of 5000 Hz were used. The free-induction decay was digitized into 30K or 34K data points. Typically 8 or 16 transients were coadded and a 15-s repetition time was used. The integrated intensity of the resonance for the C(2) protons was measured relative to that for the C(4) protons.
Results and Discussion Stepwise macroconstants for the protonation processes which take place in the basic-neutral pD region in D 2 0 are reported in Table I for seven imidazole-containing compounds. The carboxyl groups of histidylglycine, histidine, imidazole-4-acetic acid, and L-0-imidazolelactic acid are completely ionized above pH = 5, where protonation of the imidazole and amino groups takes place. Thus, protonation of the carboxylate groups is neglected here. Rate constants for substitution of deuterium for the C(2) hydrogen of those microspecies for which equilibrium constants are known are reported in Table 11. The values used for pKw(D,o) in eqs 11 and 12 were 14.385 (313 K), 14.103 (323 K), and 13.848 (333 K). Values reported for the microscopic rate constant k++ are average values from 1 to 4 series of kinetic experiments at each temperature, with each series of experiments consisting of measurements on solutions at 4-6 different pD values. Values reported for the rate constant k+ were obtained from 1 to 2 series of experiments at each temperature, with each series of experiments consisting of measurements on solutions at three or more pD values. In Figure 3, log (k++kkm)is plotted versus log kkm for histidylglycine, histidine, and histamine, and log (k+K)is plotted versus
4164 The Journal of Physical Chemistry, Vol. 95, No. 12, 1991 TABLE 11: C(2) Proton-Deuterium Exchrnge Rate Coastrnts for (Micro)npecies Where tbe Respective Equilibrium Constants Could Be Dctmnincd DireCtivOb 313 K 323 K 333 K
k++
k+
k++
k+
histidylglycine 6.79 7.23 histidine 6.67 7.04 histamine 6.56 6.96 N-acetylhistidine 6.11 6.58 imidazole-4-acetic acid 5.96 6.40 L-8-imidazolelactic acid 5.96 6.35 4-mcthylimidazole 5.74 6.19
k++ 7.63 7.51 7.38
k+
histidylglycine histidine histamine
313 K 13.20 13.26 13.33
323 K 13.44 13.51 13.58
TABLE I V MicroscoDic Rate Constant@
313 K histidylglycine histidine histamine
6.79 6.67 6.56
6.17 6.07 5.95
323 K 7.23 7.04 6.91
6.60 6.49 6.38
333 K 7.63 7.51 7.38
7.03 6.94 6.84
"Reported as log k values. Rate constant units are dm3 mol-l h-I. 7.02 6.83 6.8 1 6.64
#Rate constants are reported as log k values. Rate constant units are dm3 mol-' h-I. bEstimated uncertainty in the rate constants is 0.02-0.05 log k units. TABLE 111: Loparitbmic Values of tbe Products knk" Determined from 11
Noszil and Rabenstein
TABLE V Protonation Microconstants in D20(in log k Units)
histidylglycine
histidine
As
kN k" k2l
khm kN k" k2l
333 K 13.68 13.74 13.80
histamine
:I k" k2l
kb
313 K 7.65 7.03 6.75 6.13 9.30 7.19 8.35 6.24 10.13 7.38 9.18 6.43
323 K 7.46 6.84 6.54 5.92 9.12 7.02 8.22 6.12 9.88 7.20 8.99 6.31
333 K 7.22 6.65 6.33 5.76 8.79 6.80 7.89 5.90 9.55 6.96 8.69 6.10
TABLE VI: Protonation Microconstants at 298 K in D,O
log log log log
13.0-
12.8 5
7
6
8
log (equilibrium constant)
Figure 3. Plot of log (k++kF) and log (k+K) values versus log k$ and log K protonation constants. Points from left to right are for histidyl-
glycine, histidine, histamine, acetylhistidine, imidazole-4-acetic acid, L-8-imidazolelactic acid, and 4-methylimidazole at each temperature. See text for further details.
log K for N-acetylhistidine, imidazole-4-acetic acid, L-8imidazolelactic acid, and 4-methylimidazole. log (k++khm)and log (k+K)were calculated from the and K values in Table I and the k++ and k+ values in Table 11. The straight lines through the experimental points at each temperature are given by the following equations, which were calculated by the standard linear regression method.
kk
+ 10.580 Y323 K = 0.390X + 10.772 Y313K = 0.373X
Y333 K
0.389X
+ 11.097
(13) (14) (15)
where Y and X represent the log (rate constant X equilibrium constant) and log (equilibrium constant) values, respectively. The correlation coefficients for the data plotted in Figure 3 are 0.998 (313 K), 0.999 (323 K), and 0.997 (333 K). Equations 13, 14, and 15 are the key relationships for determination of the microscopic rate constants k,,+ and the microscopic equilibrium constant klm, for the minor protonation isomers of histidine, histamine, and histidylglycine from the experimentally determined values for the products ko+klm(eq 1 1). The experimentally determined values
kN kIm
k2l khm
histidylglycine 8.02 7.35 7.10 6.43
histidine 9.70 7.53 8.77 6.60
histamine 10.63 7.74 9.6 1 6.72
for log (k,,+k") are listed in Table 111. Using these values and eq 13-15, we calculated the microscopic rate constant k,,+ and the microscopic equilibrium constant k". The values obtained for kW are reported in Table IV and those for k" are reported in Table V. Having the value for klm, the entire sets of microconstants were calculated for histidylglycine, histidine, and histamine. The values of k k were calculated by using eq 6 and the respective kIm and constants. For histamine and histidine, microconstants kN and $j" are essentially equal to their Kl and K2values, as discussed above. However, for histidylglycine, the same simplification is not valid because the amino and imidazole basicities are more similar. Thus, for histidylglycine, kN and were also calculated by using eqs 5 and 6 and the values for k" and the macroconstants. The microconstants are reported in Table V. Microconstants a t 298 K, where the kinetics of C(2) hydrogen-deuterium exchange are too slow to measure, were obtained by extrapolation of the microconstants determined a t 313, 323, and 333 K. For histidylglycine, all microconstants were extrap olated to 298 K for histidine and histamine only k" and k k were obtained at 298 K by extrapolation since the experimentally determined macroconstants KI and K2are essentially equal to kN and khm. The microconstants at 298 K are listed in Table VI. The validity of the extrapolation procedure for obtaining microconstants at 298 K can be tested by calculating macroconstants for histidylglycine using the extrapolated values for the microconstants and eqs 5 and 6. For histidylglycine, all four of the microconstants were obtained by extrapolation, and none is insignificant when compared to the appropriate macroconstant. The values calculated for log K Iand log P2 by using the microconstants in Table VI are 8.09 and 14.48 as compared to values of 8.10 and 14.45 determined directly by potentiometry. The agreement indicates the validity of the method. The protonation microconstants in Table VI are for histidylglycine, histidine, and histamine in D20solution. Protonation microconstants can be calculated for these compounds in H20 solution by using intramolecular interactivity parameters obtained from the results in Table VI and the macroconstants measured at 298 K in H20solution. The macroconstants are as follows: histidylglycine, log K, = 7.89, log K2 = 6.23; histidine, log KI =
kk
J . Phys. Chem. 1991,95,4765-4772
TABLE MI:
ROtOartiOn MiCmwmts It 298
histidylglycine log kN log kIm 108 krn log kk
7.77 7.27 6.85 6.35
histidine 9.58 7.40 8.65 6.47
K in H20 histamine 10.32 7.64 9.30 6.62
9.58, log K2 = 6.47; histamine, log KI= 10.32, log K2 = 6.62. The microconstants calculated for H 2 0 at 298 K are reported in Table VII. Arrhenius plots of the microscopic rate constants k++ and ko+ provided the microscopic activation energies E++and Eo+ for C(2) hydrogen-deuterium substitution in the N+,Im+ and N,Im+ protonation states, respectively. The values obtained for E++and Eo+, respectively, are 84 and 86 kJ/mol for histidylglycine, 83 and 90 W/mol for histidine, and 82 and 90 kJ/mol for histamine.
Conclusions The interactivity parameter is approximately 0.9 log k units for histidine and histidylglycine. It is slightly larger and more temperature dependent for histamine, apparently due to the lack of a second, bulkier substituent (e.g., carboxylate or peptide group) that could also interact with the imidazole. When one of the histamine a-hydrogens is replaced by the -COO- or -CONHC H 2 - C 0 0 - groups, the adjacent amino basicity decreases by approximately 0.7 and 2.5 log k units, respectively, whereas the basicity of the more remote imidazole groups is diminished by only 0.2 and 0.4 units. Consequently, the amino/imidazole basicity ratio increases dramatically in the order histidylglycine C histidine C histamine. The method used in this study provides overall basicity data for the imidazole ring. However, all the imidazole constants are a composite of the N I and N3 basicities. Tanokura has determined relative values, for the N3 and N I basicities by studying N3 and N I methyl derivatives."J2 For histidine, he obtained a N,/N3 basicity ratio of 4 at 37 'C and I = 0.1. However, this data is valid only when the a-amino group is protonated. When the a-amino is blocked by acetylation, the Nl/N3 ratio is significantly lower (1.6-2.5 in different derivatives), and there are no data when the amino group is in its unmodified, neutral form. Consequently, no individual interactivity parameters can be determined for the
4765
amino-imidazole N I and the amino-imidazole No interaction without further experiments. The difference between the log k++ and log kD+ microscopic rate constants measures the effect of the amino protonation on the C(2) hydrogen-deuterium substitution. This C(2) amino interactivity parameter is approximately 0.6 in all cases but has an opposite sign from the imidazole amino interactivity. Thus, when the amino group is protonated, the imidazole N basicity is decreased, however, the C(2)deprotonation rate increased. Both phenomena are apparently a consequence of the decrease in electron density on the imidazole ring upon protonation of the amino group. For simplicity, we investigated relatively small molecules in the present study. Nevertheless, these data can also be of practical use in larger biopolymers. N-Acetylhistidine is a model for Cterminal histidine residues in peptides, while the histidyl moiety in histidylglycine is an N-terminal histidine. By taking into account the basicity differences between the corresponding microforms of histidylglycine and histidine and combining this value with the log K value for N-acetylhistidine, we estimate log k 7.2 in H 2 0 at 298 K and I = 2.0 M dm-3 for the imidazole of a histidyl residue in a peptide chain. By an analogous procedure, the rate constants for C(2)-hydrogen substitution for histidyl residues in peptides is log k 6.2, where k has units of dm3 mol-' h-' in D 2 0 at 313 K and I = 2.0 M dm-). These values may be modified in most peptides and proteins by intra- and intermolecular interactions. The microscopic activation energy values have an estimated uncertainty of 2-4 kJ mol-'. However, these parameters show unambiguously that the rupture of the C(2)-proton bond requires a few kJ mol-' less activation energy when both the imidazole and the amino groups are protonated. Acknowledgment. This research was supported by National Institutes of Health Grant AI24216. B.N. gratefully acknowledges a fellowship from the Soros Foundation. The NMR instrumentation was supported in part by BRSG 2 SO7 RR 07010-20 awarded by Biomedical Research Resources, National Institutes of Health, and B.P. America. Rdstry No. H-His-Gly-OH, 2578-58-7; H-His-OH, 71-00-1; AcHis-OH, 2497-02-1; 4-methylimidazole, 822-36-6; histamine dihydrochloride, 56-92-8; imidazole-4-acetic acid, 645-65-8; L-j3-imidazolelactic acid, 14403-45-3.
=
Reactions of Os+, Ar+, Ne+, and He+ with SICI,: Thermochemistry of SiCi,+ ( x = 1-3) Ellen R. Fisher and P.B. Armentrout**t Department of Chemistry, University of Utah, Salt Lake City, Utah 84112 (Received: December 12, 1990) Reactions of 02+, Ar+, Ne+, and He+ with SiC14 are studied by using guided ion beam mass spectrometry. All reactions are found to be fairly efficient with thermal energy rate constants that exceed 66% of the collision rate. The major products and Ar+ reactions are SiC14+and SiC13+,while for the Ne+ and He+ reactions, the major product is observed in the 02+ SiCI'. Thermochemistry derived from these SiCI, systems includes the determination of AfH(SiC13+)= 99.8 f 1.6 kcal/mol system, and ArH(SiCI2+)= 184.9 f 2.6 kcal/mol and AfH(SiCI+)= 203.9 and AfH(OSiC13+)< 97 kcal/mol from the 02+ f 2.5 kcal/mol from the Ar+ system. The ionization energies IE(SiC13) = 7.65 f 0.15 eV, IE(SiCI2) = 9.81 f 0.10 eV, and IE(SiC1) = 6.79 f 0.24 eV are also derived after consideration of other literature thermochemistry.
Introduction In the fabrication of microelectronic devices, chlorosilanes are used extensively in chemical vapor deposition (CVD) and plasma-enhanced CVD systems to deposit silicon layers.' Chlorine-based plasmas that form silicon chloride ions and radicals are also often used to etch such layers.lc.2 A detailed understanding of the chemical mechanisms involved in these plasmas 'Camille and Henry Dreyfus Teacher-Scholar. 1987-1992.
0022-3654/9l/2095-4765$02.50/0
can provide insight into the most important physical parameters of a plasma or CVD reactor. Thus the investigation of the reactivity,) structure: and thermochemistry>* of silicon chloride (1) (a) Inspektor-Koren, A. Sur/. Coat. Technol. 1987,33,3I . (b) Ban, V. J . Electrochem. SOC.lWS,122,1389. (c) Much, J. A.; Ha,D. W. ACS Symp. Ser. 1983, 219, 215. (2) (a) Schwartz, G. C.; Schaible, P. M. J . Voc. Scl. Technol. 1979, 16, 410. (b) Wormhoudt, J.; Stanton, A. C.; Richards, A. D.; Swain, H. H. J. Appl. Phys. 1981,61, 142.
63 1991 American Chemical Society
