Fourier transform infrared spectra of the L intermediate of light-adapted bacteriorhodopsin were examined for recombinant proteins with amino acid substitutions at Thr46 and Asp96. Two O-H stretching vibrational bands of water, at 3607 and 3577 cm-1, change into stronger H-bonding states in L of the wild type. Thr46-->Val substitution abolished these bands in spite of the fact that [3-18O]threonine-labeling did not shift them, indicating that they correspond to coordination of the water with Thr46. The two water bands were restored, although with changed frequencies, by an additional Asp96-->Asn substitution in Thr46-->Val/Asp96-->Asn. A single Asp96-->Asn substitution abolished the 3607 cm-1 band. Thus, Asp96 also takes part in structural changes in water. The perturbations of these water molecules in the L intermediate displayed a weak correlation with the ratio of intensity change in the two vibrational bands of the Schiff base mode at 1312 and 1301 cm-1 and the rate for the deprotonation of the Schiff base at the L-to-M reaction of the photocycle. We find, therefore, that the water molecules in the cytoplasmic Asp96-Thr46 domain, which comprises the site of proton uptake after formation of the M intermediate, undergo structural changes in the L intermediate already. These changes are transmitted to the extracellular domain and affect interaction of the Schiff base with Asp85, that is far removed from this region.

[reaction: see text] By choosing the right substituents either highly functionalized unusual four-membered ring amino acids or the isomeric pipecolic acid derivatives are obtained in enantiomerically pure form. Starting material is a linear allene-containing amino acid that has been resolved via biocatalysis.

Abstraen û>(2,3-ButadienyI)lactams react with aryl iodides in the presence of n-flu^NCl, K2COj and a catalytic amount of Pd(PPh3)4 to give bicyclic enamides resulting from attack of nitrogen on the central carbon and transfer of the aryl group to the terminal carbon atom of the aliene.

A route for the synthesis of enantiopure allene-substituted lactams has been developed. The key-step involves the copper(I) mediated S N 2' substitution of propargylic tosylates by a (S)pyroglutamic acid derived organozinc reagent. Pd-catalysed reaction of these allenes with iodobenzene afforded enantiopure bicyclic enamides. Furthermore the unexpected formation of an interesting diene is reported.

[3‐18O]‐L‐serine, [3‐13C]‐L‐serine, [3‐18O]‐L‐threonine, [3,4‐13C2]‐L‐threonine and [3‐2H]‐L‐threonine are prepared from simple commercially available, isotopically enriched starting materials like H218O, [13C]‐paraformaldehyde, [13C2]‐acetaldehyde and [1‐2H]‐acetaldehyde. The introduction of the side chain is based on the reaction of the anion of the bislactimether of cyclo‐(D‐Val‐Gly) with a suitable reagent. For serine this is isotopically labelled benzylchloromethylether, whereas for threonine labelled acetaldehyde is used in combination with chlorotitaniumtris[diethylamide], introducing both stereocentres in one single step. The isotopomers of serine and threonine are obtained on the gram scale in good yields and high enantiomeric and diasteriomeric excesses. New syntheses for [18O]‐benzylalcohol and isotopically enriched benzylchloromethylether are reported. Following the presented synthetic scheme these amino acids can be labelled at any position or at any combination of positions.

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