Isolation and characterization of distinct bioactive forms of LH from male buffalo pituitaries: differences localized to their alpha subunits.
The isolation of highly purified forms of pituitary LH from Egyptian male (Nile) buffaloes is described. The total LH content (receptor binding activity) which was approximately 30 to 50 fold higher than FSH in the pituitary could be divided into three pools based upon fractionation patterns on a cation exchanger. The acidic fraction which also contained FSH was not purified to homogeneity. A basic fraction (bu-LH-2; 300 mg/kg anterior pituitary) and a very basic fraction (bu-LH-3; 80 mg/kg) were both highly purified and free of FSH activity as tested by specific FSH receptor and immunoassays. The basic buffalo LH fraction, bu-LH-2, was as active as highly purified ovine LH (oLH). The most basic form of buffalo LH, bu-LH-3, was, however, about twice as active as highly purified oLH in the in vitro bioassay using mouse Leydig tumour (MA-10) cells. In a receptor binding assay employing 125I-labelled buffalo LH (bu-LH-3) and porcine testicular membranes, the affinity of bu-LH-3 was about five times higher than purified oLH. The M(r) of both forms of purified buffalo LH and subunits was similar to that of oLH. Amino acid composition of buffalo LH was also very similar to oLH except for small differences. Fractionation by fast protein liquid chromatography on Mono-Q columns revealed further evidence of microheterogeneity in each of the pools of buffalo LH with bu-LH-3 exhibiting a predominant single component. By reverse-phase high-pressure liquid chromatography analysis we have localized differences in the two purified isoforms of male buffalo LH to the alpha subunit. It is suggested that differences in biological potencies could be due to variations in terminal glycosylation and/or differences in branching of this subunit which is known to be important for signal transduction.
Sairam
M R
MR
Reproduction Research Laboratory, Clinical Research Institute of Montreal, Québec, Canada.
Zaky
A A
AA
Hassan
A A
AA
D000596:Amino Acids / Q000032:analysis; D000818:Animals; D001681:Biological Assay; D002020:Buffaloes / Q000378:metabolism*; D002851:Chromatography, High Pressure Liquid; D004591:Electrophoresis, Polyacrylamide Gel; D005640:Follicle Stimulating Hormone / Q000378:metabolism; D015292:Glycoprotein Hormones, alpha Subunit / Q000032:analysis; D007986:Luteinizing Hormone / Q000032:analysis* / Q000737:chemistry / Q000378:metabolism; D008297:Male; D010902:Pituitary Gland / Q000737:chemistry*; D011863:Radioimmunoassay; D011869:Radioligand Assay; D012756:Sheep
D000596:Amino Acids; D015292:Glycoprotein Hormones, alpha Subunit; D007986:Luteinizing Hormone; D005640:Follicle Stimulating Hormone