Both trans-syn cyclobutane-type photodimers of 2'-deoxyuridylyl (3'-5') thymidine (dUpdT) were formed by deamination of the corresponding trans-syn cyclobutane photodimers of 2'-deoxycytidylyl (3'-5') thymidine (dCpdT) and were examined by 1H-, 13C-, and 31P-nmr spectroscopy. One- and two-dimensional nmr experiments provided a nearly complete assignment of the 1H, 13C, and 31P resonances. Scalar and nuclear Overhauser effect contacts were used to determine the conformation of the deoxyribose rings, exocyclic bonds, cyclobutane rings, and glycosidic linkages. Isomer I (S-type class; CB-; SYN-ANTI) and isomer II (N-type class; CB+; ANTI-SYN) exhibit markedly different conformational features. 31P chemical shifts show that the relative flexibility is dUpdT > isomer II > isomer I. The conformations of these species are very similar to those of other previously examined trans-syn photodimers. Among bipyrimidine photodimers of a given diastereomeric form (i.e., trans-syn I or II), the nmr-derived conformational parameters are nearly invariant, regardless of base substitution pattern. This contrasts with the substituent-dependent variation of cyclobutane ring conformation observed by Kim et al. (Biopolymers, 1993, Vol. 33, pp. 713-721) for an analogous series of cis-syn photodimers. Steric crowding of cyclobutane ring substituents is offered as an explanation for the difference in substituent effects between the families of cis-syn and trans-syn photodimers.