Piv-Proψ[CH₂–NH–O]Gly–NHⁱPr

The pseudodipeptide, (S)-N-iso­propyl {[N-(pivaloyl)­pyrrol­idin-2-yl]­methyl­amino­oxy}acet­amide, C₁₅H₂₉N₃O₃, adopts a global extended conformation with the hydroxy­l­amine group in the g⁺/g⁻ structure. The C-terminal amide NH interacts intramolecularly with the hydroxy­lamine O atom. Both NH bonds of each mol­ecule are hydrogen bonded to the C-­terminal amide carbonyl of a neighbouring mol­ecule.

S1. Comment

The so-called reduced peptide group Cα-CH₂—NH—Cα is often used to mimic the transient state Cα-C(OH)₂—NH—Cα of the peptide group during enzymatic cleavage of the amide bond (Epps et al., 1988; Sawyer, Pals, Mao, Maggiora et al., 1988; Sawyer, Pals, Mao, Staples et al., 1988; Kaltenbronn et al., 1990; Gante, 1994). However, the pK_a of this amide surrogate is about neutrality and may depend on the environment, so that at least a part of the molecules may be protonated into Cα-CH₂—N⁺H₂—Cα at the physiological pH (Aumelas et al., 1987; Vanderesse et al., 1998). Contrary to the neutral form, the ionic form is a proton donor capable of strong hydrogen bonds with nucleophiles, that may induce particular folded structures (Vanderesse et al., 1998) or interaction modes in intermolecular interactions. We have recently proposed two reduced amide surrogates having the N—N or N—O fragment that decreases the pK_a to such a value that it is not protonated at the physiological pH (Vanderesse et al., 1998; Thévenet et al., 2000). Here we report the crystal molecular structure of a pseudodipeptide containing the methyleneaminoxy link, (I). \sch

The three-dimensional structure (Table 1) shows that the C6—C10—N2—O2—C11 hydroxylamine fragment assumes a skew conformation, so that the C6···C11 (Cα···Cα) distance of 3.682 (4) Å is a little bit shorter than the corresponding distance of 3.81 Å for a peptide group (Benedetti, 1977). The molecule assumes a globally extended conformation (Fig. 1) in which the C-terminal NH is intramolecularly hydrogen-like bonded to the hydroxylamine oxygen (Table 2), exactly as in chloroform solution (Thévenet et al., 2000). Although it is out of the usual criteria defining hydrogen bonds (Baker & Hubbard, 1984), such a bent interaction has already been encountered in various crystal molecular structures of modified peptides (Toniolo et al., 1989; Aubry et al., 1994; Crisma et al., 1999). Molecules are held in files along the x axis by a double interaction involving both NH groups of a given molecule and the C-terminal amide carbonyl of another molecule (Table 2).

S2. Experimental

The pseudodipeptide was obtained by coupling NH₂—O—CH₂—CO—NHⁱPr to the Boc-L-Pro-H aldehyde, with subsequent ^tBuCO (Piv) for tert-butyloxycarbonyl (Boc) substitution, and reduction of the oxime by NaBH₃CN. The commercially available Boc-NH—O—CH₂—COOH (382 mg, 2 mmol) and N-methylmorpholine (NMM) (0.22 ml, 2 mmol) in tetrahydrofuran (THF) (20 ml) were treated dropwise under stirring with isobutylchloroformate (0.26 ml, 2 mmol) diluted in THF (2 ml) at 255 K, and stirring was maintained for 15 min. Isopropylamine (0.17 ml, 2 mmol) in THF (2 ml) was added dropwise, and the temperature was allowed to reach room temperature. The mixture was stirred overnight. NMM hydrochloride was filtered off and the solvent was evaporated. Boc-NH—O—CH₂—CO—NHⁱPr was purified by silica gel chromatography with ethyl acetate/petroleum ether (60/40, v/v) as eluent (Rf = 0.58, 348 mg, 1.5 mmol, yield = 75%). The Boc group was quantitatively eliminated with trifluoroacetic acid (TFA) in dichloromethane (DCM) (40/60), and TFA·NH₂—O—CH₂—CO—NHⁱPr was recovered by lyophilization from an aqueous solution. TFA·NH₂—O—CH₂—CO—NHⁱPr (369 mg, 1.5 mmol) in ethanol (20 ml) was treated with NMM (0.16 ml, 1.5 mmol) and coupled to Boc-L-Pro-H (597 mg, 3 mmol), obtained from Boc-L-Pro-OH (Fehrentz & Castro, 1983), in the presence of sodium acetate (492 mg, 6 mmol) and molecular sieves. The mixture was stirred at room temperature overnight to give Boc-Proψ[CH=N—O]Gly-NHⁱPr which was purified by silica gel chromatography with ethyl acetate/petroleum ether (70/30, v/v) (Rf = 0.53, 385 mg, 1.23 mmol, yield = 82%). The Boc group was quantitatively eliminated with TFA in DCM (40/60), and the resulting TFA·H-Proψ[CH═N—O]Gly-NHⁱPr was obtained by lyophilization from an aqueous solution. TFA·H-Proψ[CH═N—O]Gly-NHⁱPr (402 mg, 1.23 mmol) was dissolved in chloroform (20 ml), the solution was cooled to 273 K, and diisopropylethylamine (0.42 ml, 2.46 mmol) and pivaloyl chloride (0.23 ml, 1.85 mmol) were successively added dropwise. The mixture was stirred at 273 K for 2 h to give Piv-Proψ[CH═N—O]Gly-NHⁱPr which was purified by silica gel chromatography with ethanol/ethyl acetate/petroleum ether (10/60/30, v/v/v) (Rf = 0.67, 245 mg, 0.82 mmol, yield = 67%). Piv-Proψ[CH═N—O]Gly-NHⁱPr (245 mg, 0.82 mmol) was dissolved in methanol (10 ml) and NaBH₃CN (515 mg, 8.2 mmol) was added under stirring at room temperature in 8 portions over 96 h while the pH was adjusted to 3 with acetic acid (acid-base indicator: methyl orange). The solution was poured into 5 ml of water saturated with K₂CO₃. The solution was repeatedly extracted with DCM (5 x 5 ml). The organic phases were combined and washed three times with 5% aqueous NaHCO₃ and three times with brine. Piv-Proψ[CH₂—NH—O]Gly-NHⁱPr was purified by silica gel chromatography with ethanol/ethyl acetate/petroleum ether (10/60/30, v/v/v) as eluent (Rf = 0.45, 125 mg, 0.42 mmol, yield = 51%). Single crystals were obtained by slow evaporation of a DCM solution.

S3. Refinement

The absolute stereochemistry of the title compound was assumed from Boc-L-Pro-OH purchased from Neosystem corporation (Strasbourg, France). The position of H atoms attached to N atoms were located from a difference map and the N—H bond distance was restrained to 1.03 (1) Å (Taylor & Kennard, 1983). H atoms connected to carbon were placed at calculated positions using a riding model. All hydrogen atoms have isotropic displacement displacement parameters fixed at 1.3 times that of the parent atom.

Computing details

Data collection: CAD-4 Software (Enraf-Nonius, 1989); cell refinement: CAD-4 Software; data reduction: maXus (Mackay et al., 1999); program(s) used to solve structure: SIR92 (Altomare et al., 1994); program(s) used to refine structure: SHELXL97 (Sheldrick, 1997); molecular graphics: maXus.

Figures

Fig. 1. The molecular structure of the pseudodipeptide with the atom-numbering scheme and 25% probability displacement ellipsoids. H atoms except those of the NH groups are omitted for clarity.

(S)—N-(tert-butylcarbonyl)-2-[[O-(N-isopropylacetamidyl)] -hydroxylaminomethylene]-pyrrolidine

Crystal data

C15H29N3O3	Dx = 1.169 Mg m^−3
Mr = 299.41	Cu Kα radiation, λ = 1.54060 Å
Orthorhombic, P212121	Cell parameters from 25 reflections
a = 5.3860 (5) Å	θ = 9.7–26.2°
b = 11.470 (2) Å	µ = 0.66 mm^−1
c = 27.539 (3) Å	T = 293 K
V = 1701.3 (4) Å^3	Prismatic, colorless
Z = 4	0.6 × 0.1 × 0.1 mm
F(000) = 656

Data collection

Nonius Mach3 diffractometer	Rint = 0.000
Radiation source: Nonius FR591 rotating Cu anode	θmax = 69.7°, θmin = 3.2°
Graphite monochromator	h = 0→6
ω/2θ scans	k = 0→13
1877 measured reflections	l = 0→33
1877 independent reflections	2 standard reflections every 60 min
1702 reflections with I > 2σ(I)	intensity decay: 3.7%

Refinement

Refinement on F^2	Secondary atom site location: difference Fourier map
Least-squares matrix: full	Hydrogen site location: inferred from neighbouring sites
R[F^2 > 2σ(F^2)] = 0.054	H atoms treated by a mixture of independent and constrained refinement
wR(F^2) = 0.157	w = 1/[σ^2(Fo^2) + (0.112P)^2 + 0.304P] where P = (Fo^2 + 2Fc^2)/3
S = 1.06	(Δ/σ)max = 0.019
1877 reflections	Δρmax = 0.28 e Å^−3
197 parameters	Δρmin = −0.33 e Å^−3
2 restraints	Extinction correction: SHELXL97 (Sheldrick, 1997), Fc^*=kFc[1+0.001xFc^2λ^3/sin(2θ)]^-1/4
Primary atom site location: structure-invariant direct methods	Extinction coefficient: 0.0125 (16)

Special details

Geometry. All e.s.d.'s (except the e.s.d. in the dihedral angle between two l.s. planes) are estimated using the full covariance matrix. The cell e.s.d.'s are taken into account individually in the estimation of e.s.d.'s in distances, angles and torsion angles; correlations between e.s.d.'s in cell parameters are only used when they are defined by crystal symmetry. An approximate (isotropic) treatment of cell e.s.d.'s is used for estimating e.s.d.'s involving l.s. planes.

Refinement. Refinement of F² against ALL reflections. The weighted R-factor wR and goodness of fit S are based on F², conventional R-factors R are based on F, with F set to zero for negative F². The threshold expression of F² > σ(F²) is used only for calculating R-factors(gt) etc. and is not relevant to the choice of reflections for refinement. R-factors based on F² are statistically about twice as large as those based on F, and R- factors based on ALL data will be even larger.

Fractional atomic coordinates and isotropic or equivalent isotropic displacement parameters (Ų)

	x	y	z	Uiso*/Ueq
O2	0.3926 (4)	0.23774 (17)	0.07053 (7)	0.0442 (5)
O3	0.9164 (4)	0.4192 (2)	0.04322 (9)	0.0580 (6)
O1	0.2749 (5)	0.25799 (18)	0.25725 (7)	0.0562 (6)
N1	0.2908 (4)	0.08643 (17)	0.21961 (8)	0.0383 (5)
N3	0.5056 (5)	0.4479 (2)	0.02998 (10)	0.0468 (6)
H3	0.328 (3)	0.416 (3)	0.0294 (14)	0.061*
N2	0.3164 (5)	0.29575 (19)	0.11501 (8)	0.0453 (6)
H2	0.178 (5)	0.348 (3)	0.1018 (12)	0.059*
C12	0.7006 (5)	0.3856 (2)	0.04434 (10)	0.0431 (6)
C6	0.4081 (5)	0.1432 (2)	0.17694 (10)	0.0397 (6)
H6	0.5377	0.1974	0.1876	0.052*
C5	0.2316 (5)	0.1526 (2)	0.25858 (9)	0.0389 (6)
C10	0.2102 (6)	0.2081 (2)	0.14761 (10)	0.0431 (6)
H10A	0.1162	0.1523	0.1286	0.056*
H10B	0.0961	0.2461	0.1698	0.056*
C13	0.5235 (6)	0.5661 (3)	0.01026 (12)	0.0530 (8)
H13	0.6978	0.5904	0.0090	0.069*
C1	0.1008 (5)	0.1012 (3)	0.30359 (11)	0.0475 (7)
C9	0.2741 (7)	−0.0412 (2)	0.21346 (11)	0.0506 (7)
H9A	0.1070	−0.0689	0.2198	0.066*
H9B	0.3889	−0.0813	0.2349	0.066*
C11	0.6451 (6)	0.2631 (3)	0.06192 (13)	0.0534 (8)
H11A	0.7075	0.2083	0.0380	0.069*
H11B	0.7366	0.2499	0.0918	0.069*
C7	0.5233 (7)	0.0403 (3)	0.15064 (12)	0.0575 (8)
H7A	0.5368	0.0556	0.1161	0.075*
H7B	0.6870	0.0229	0.1634	0.075*
C4	0.1954 (8)	−0.0178 (3)	0.32134 (14)	0.0700 (10)
H4A	0.3667	−0.0113	0.3305	0.091*
H4B	0.1792	−0.0741	0.2957	0.091*
H4C	0.0992	−0.0424	0.3488	0.091*
C8	0.3446 (8)	−0.0588 (3)	0.16046 (13)	0.0613 (9)
H8A	0.4242	−0.1338	0.1556	0.080*
H8B	0.1999	−0.0537	0.1396	0.080*
C3	0.1355 (9)	0.1880 (4)	0.34519 (13)	0.0760 (12)
H3A	0.3088	0.1944	0.3529	0.099*
H3B	0.0463	0.1613	0.3732	0.099*
H3C	0.0731	0.2629	0.3356	0.099*
C14	0.3775 (11)	0.6485 (3)	0.04224 (18)	0.0863 (13)
H14A	0.4540	0.6527	0.0737	0.112*
H14B	0.2104	0.6204	0.0456	0.112*
H14C	0.3753	0.7246	0.0278	0.112*
C2	−0.1714 (6)	0.0938 (6)	0.29024 (17)	0.0955 (18)
H2A	−0.1928	0.0384	0.2644	0.124*
H2B	−0.2286	0.1689	0.2798	0.124*
H2C	−0.2653	0.0692	0.3180	0.124*
C15	0.416 (2)	0.5665 (4)	−0.03952 (14)	0.132 (3)
H15A	0.5136	0.5174	−0.0603	0.171*
H15B	0.4160	0.6446	−0.0520	0.171*
H15C	0.2486	0.5378	−0.0383	0.171*

Atomic displacement parameters (Ų)

	U^11	U^22	U^33	U^12	U^13	U^23
O2	0.0472 (11)	0.0398 (9)	0.0454 (10)	−0.0002 (9)	0.0024 (9)	0.0028 (8)
O3	0.0357 (10)	0.0630 (13)	0.0752 (14)	−0.0026 (11)	0.0017 (10)	0.0116 (12)
O1	0.0694 (15)	0.0398 (10)	0.0594 (11)	−0.0068 (12)	0.0066 (11)	−0.0005 (8)
N1	0.0369 (11)	0.0300 (9)	0.0482 (11)	−0.0010 (10)	0.0008 (10)	0.0092 (8)
N3	0.0353 (12)	0.0434 (12)	0.0618 (14)	−0.0006 (11)	0.0025 (11)	0.0166 (11)
N2	0.0566 (14)	0.0330 (10)	0.0465 (12)	0.0043 (12)	0.0063 (11)	0.0051 (9)
C12	0.0388 (13)	0.0448 (14)	0.0458 (13)	0.0017 (13)	0.0065 (12)	0.0052 (11)
C6	0.0325 (12)	0.0371 (12)	0.0494 (14)	0.0023 (12)	0.0012 (11)	0.0086 (11)
C5	0.0298 (12)	0.0390 (13)	0.0479 (13)	−0.0029 (12)	−0.0021 (11)	0.0077 (10)
C10	0.0400 (13)	0.0396 (13)	0.0496 (14)	0.0069 (13)	0.0042 (12)	0.0109 (11)
C13	0.0462 (15)	0.0431 (15)	0.0696 (18)	−0.0064 (15)	0.0037 (14)	0.0191 (13)
C1	0.0316 (13)	0.0607 (17)	0.0501 (14)	−0.0036 (14)	0.0017 (11)	0.0094 (13)
C9	0.0568 (18)	0.0307 (12)	0.0643 (16)	−0.0012 (13)	−0.0017 (15)	0.0071 (11)
C11	0.0420 (15)	0.0473 (16)	0.0711 (18)	0.0086 (14)	0.0122 (14)	0.0144 (14)
C7	0.0558 (18)	0.0535 (17)	0.0630 (17)	0.0232 (16)	0.0121 (15)	0.0089 (14)
C4	0.068 (2)	0.070 (2)	0.072 (2)	−0.009 (2)	0.0076 (19)	0.0344 (18)
C8	0.077 (2)	0.0377 (14)	0.0690 (18)	0.0118 (16)	0.0012 (19)	−0.0010 (13)
C3	0.085 (3)	0.090 (3)	0.0534 (17)	−0.006 (3)	0.0084 (19)	0.0026 (17)
C14	0.105 (4)	0.0549 (19)	0.099 (3)	0.008 (3)	−0.004 (3)	−0.005 (2)
C2	0.0302 (16)	0.177 (6)	0.080 (2)	−0.011 (3)	0.0022 (16)	0.015 (3)
C15	0.267 (9)	0.074 (2)	0.054 (2)	−0.014 (5)	−0.008 (4)	0.0246 (19)

Geometric parameters (Å, º)

O2—C11	1.411 (4)	C6—C7	1.518 (4)
O2—N2	1.453 (3)	C6—C10	1.531 (4)
O3—C12	1.225 (4)	C5—C1	1.543 (4)
O1—C5	1.232 (4)	C13—C15	1.489 (6)
N1—C5	1.353 (4)	C13—C14	1.513 (6)
N1—C9	1.477 (3)	C1—C2	1.514 (4)
N1—C6	1.485 (3)	C1—C3	1.530 (5)
N3—C12	1.330 (4)	C1—C4	1.536 (5)
N3—C13	1.464 (4)	C9—C8	1.522 (5)
N2—C10	1.464 (3)	C7—C8	1.514 (5)
C12—C11	1.516 (4)
C11—O2—N2	108.6 (2)	N2—C10—C6	112.7 (2)
C5—N1—C9	129.3 (2)	N3—C13—C15	108.6 (3)
C5—N1—C6	118.8 (2)	N3—C13—C14	109.2 (3)
C9—N1—C6	111.7 (2)	C15—C13—C14	109.4 (4)
C12—N3—C13	123.7 (3)	C2—C1—C3	109.7 (4)
O2—N2—C10	108.2 (2)	C2—C1—C4	110.4 (4)
O3—C12—N3	124.9 (3)	C3—C1—C4	107.4 (3)
O3—C12—C11	119.1 (3)	C2—C1—C5	105.6 (3)
N3—C12—C11	115.9 (3)	C3—C1—C5	107.3 (3)
N1—C6—C7	102.2 (2)	C4—C1—C5	116.3 (3)
N1—C6—C10	109.6 (2)	N1—C9—C8	103.1 (2)
C7—C6—C10	114.3 (3)	O2—C11—C12	115.7 (2)
O1—C5—N1	118.9 (2)	C8—C7—C6	103.8 (2)
O1—C5—C1	119.1 (3)	C7—C8—C9	103.3 (3)
N1—C5—C1	122.0 (2)
C13—N3—C12—O3	0.6 (5)	C12—N3—C13—C15	121.0 (5)
C5—N1—C6—C7	161.1 (3)	C12—N3—C13—C14	−119.8 (4)
C9—N1—C6—C7	−14.6 (3)	O1—C5—C1—C2	96.9 (4)
C1—C5—N1—C6	177.8 (2)	N1—C5—C1—C2	−80.3 (4)
C5—N1—C6—C10	−77.4 (3)	O1—C5—C1—C3	−20.0 (4)
N1—C6—C10—N2	161.3 (2)	N1—C5—C1—C3	162.8 (3)
C6—C10—N2—O2	83.8 (3)	O1—C5—C1—C4	−140.3 (3)
C10—N2—O2—C11	−125.4 (2)	N1—C5—C1—C4	42.6 (4)
N2—O2—C11—C12	−73.8 (3)	C5—N1—C9—C8	174.9 (3)
O2—C11—C12—N3	−12.6 (4)	C6—N1—C9—C8	−10.0 (4)
C11—C12—N3—C13	−177.7 (3)	O3—C12—C11—O2	169.0 (3)
C9—N1—C6—C10	107.0 (3)	N1—C6—C7—C8	33.5 (3)
C9—N1—C5—O1	175.4 (3)	C10—C6—C7—C8	−84.7 (3)
C6—N1—C5—O1	0.6 (4)	C6—C7—C8—C9	−40.5 (3)
C9—N1—C5—C1	−7.5 (5)	N1—C9—C8—C7	30.7 (3)
C7—C6—C10—N2	−84.7 (3)

Hydrogen-bond geometry (Å, º)

D—H···A	D—H	H···A	D···A	D—H···A
N3—H3···O2	1.02 (1)	2.37 (4)	2.725 (3)	99 (2)
N3—H3···O3^i	1.02 (1)	2.25 (2)	3.211 (3)	156 (3)
N2—H2···O3^i	1.02 (1)	2.29 (2)	3.249 (3)	155 (3)

Symmetry code: (i) x−1, y, z.