The bacterial heterodimeric ATP‐binding cassette (ABC) multidrug exporter PatAB has a critical role in conferring antibiotic resistance in multidrug‐resistant infections by Streptococcus pneumoniae. As with other heterodimeric ABC exporters, PatAB contains two transmembrane domains that form a drug translocation pathway for efflux and two nucleotide‐binding domains that bind ATP, one of which is hydrolysed during transport. The structural and functional elements in heterodimeric ABC multidrug exporters that determine interactions with drugs and couple drug binding to nucleotide hydrolysis are not fully understood. Here, we used mass spectrometry techniques to determine the subunit stoichiometry in PatAB in our lactococcal expression system and investigate locations of drug binding using the fluorescent drug‐mimetic azido‐ethidium. Surprisingly, our analyses of azido‐ethidium‐labelled PatAB peptides point to ethidium binding in the PatA nucleotide‐binding domain, with the azido moiety crosslinked to residue Q521 in the H‐like loop of the degenerate nucleotide‐binding site. Investigation into this compound and residue’s role in nucleotide hydrolysis pointed to a reduction in the activity for a Q521A mutant and ethidium‐dependent inhibition in both mutant and wild type. Most transported drugs did not stimulate or inhibit nucleotide hydrolysis of PatAB in detergent solution or lipidic nanodiscs. However, further examples for ethidium‐like inhibition were found with propidium, novobiocin and coumermycin A1, which all inhibit nucleotide hydrolysis by a non‐competitive mechanism. These data cast light on potential mechanisms by which drugs can regulate nucleotide hydrolysis by PatAB, which might involve a novel drug binding site near the nucleotide‐binding domains.

In laboratory research on tonal coarticulation in Taiwanese, one study [H.-B, Lin, Ph.D. dissertation, Univ. of Connecticut (1988)] reported a perseveratory effect but no anticipatory effect, while another [S.-H. Peng, J. Phonetics 25, 371–400 (1997)] found a significant anticipatory effect. Peng also found tonal variation due to prosodic positions. Unlike these previous laboratory studies, this study attempts to investigate tonal coarticulation and prosodic effects on Taiwanese tones using natural conversations from the Taiwanese Spoken Corpus (Tsay and Myers, 2004), of which 56 min of recorded conversations were analyzed. Consistent with Lin, the results showed that tone is more affected by the preceding tone than by the following tone. The slope is more influenced by the preceding tone as well. As for prosodic effects, the results confirmed Peng, showing that F0 is the lowest in utterance-final position, while in other phrase-final positions it is slightly lower than in non-phrase-final position. This study thus demonstrates the results obtained in the laboratory do indeed carry over into actual conversation.