Follicle Stimulating Hormone Binding Inhibitor Produced by the Bacteria Serratia Interacts with Receptor for Follicle Stimulating Hormone in Calf Testis Membranes1

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Abstract
Bacteria of the genus Serratia, including a strain of Serratia liquefaciens isolated from contaminated porcine follicular fluid, produced an inhibitor of 125 I-human follicle-stimulating hormone (hFSH) binding to calf testis membranes in vitro. In order to evaluate its potential usefulness and significance, we undertook studies to identify the site of action of this inhibitor. Large quantities (grams) of inhibitor-containing material (SL-1) were obtained by enrichment culture techniques its chemical ocmposition was determined. Fillicle-stimulating hormone-binding inhibitory activity (FSH-BI) in SL-1 was associated with a protein-containing substance of approximately 30,000 Mr and also with larger Mr (> 300,000) forms. Preincubation studies demonstrated that binding inhibiton by SL-1 was due to effects on the membranes rather than effects on the radioligand (125 I-hFSH). Kinetics studies indicated that FSH inhibition by SL-1 was a relatively slow process that reached steady-state conditions between 23 and 25 h at 20.degree. C in contrast to FSH, which reached steady-state conditions by 12 h at 20.degree. C. Estimates of FSH-BI activity, e.g., mass required to produce a 50% inhibition of 125I-hFSH binding, varied drastically when these kinetics differences were nto taken into account. Our observations emphasize the need to establish steady-state conditions for each ligand beofre assessing mechanisms of action using Michaelis-Menton assumptions (e.g., competitive binding assays, Scatchard analyses). Binding studies conducted under steady-state conditions for FSH and SL-1 revealed that SL-1 inhibits 125I-hFSH binding via reductions in the number of available receptors for FSH without statistically signficiant effects on the apparent affinity constant (Ka) of binding. These data demonstrate that the material produced by Serratia liquifaciens is a highly potent inhibitor that interacts with the receptor for FSH.