Effects of D-amino acid substituents on degradation of LHRH analogues by proximal tubule

The luteinizing hormone-releasing hormone, LHRH, is degraded in renal proximal tubules (PT) in vivo (rat) and in vitro (rabbit) to < Glu-His (2), < Glu-His-Trp (3), and < Glu-His-Trp-Ser (4). LHRH may be cleaved by endopeptidases simultaneously at multiple bonds, or initially at Ser4-Try5 followed by carboxypeptidase hydrolysis of 4 to 3 and then 2. To distinguish between these mechanisms, [3H]LHRH analogues were incubated with rabbit renal brush-border membranes (BBM), microinfused into PT in vivo or in vitro, and products were analyzed by HPLC. [D-Ser4]LHRH was not cleaved at D-Ser4-Try5 but yielded < Glu-His-Trp-D-Ser-Tyr-Gly as the major metabolite plus 2 and 3. [D-Trp6]LHRH was cleaved by BBM and PT to 2 and 3, but not to 4. [D-Ser4, D-Trp6]LHRH was not cleaved by BBM, but was degraded to 2 by PT in vivo. Thus, D-amino acid substituents altered the expected cleavage pattern of these analogues. Thus, normally LHRH may be cleaved in PT by endopeptidase-24.11 to 2 and 4, and by angiotensin I-converting enzyme to 3, its know cleavage site