Biosynthetic 20-kilodalton methionyl-human growth hormone has diabetogenic and insulin-like activities.

Abstract

The anterior pituitary gland produces a 20-kilodalton (kDa) variant of human growth hormone (hGH) that differs from the predominant 22-kDa form of hGH in that amino acid residues 32-46 are deleted. Previous work has suggested that the 20-kDa variant possesses the full growth-promoting and lactogenic activities of 22-kDa hGH but lacks its intrinsic diabetogenic and insulin-like activities. In the present study, recombinant DNA techniques were used to prepare biosynthetic 20-kDa hGH, and some of the biological properties of the purified hGH variant were examined. The biosynthetic 20-kDa hGH variant was found to share the propensity for aggregation exhibited by its native counterpart. Moreover, like the native variant, biosynthetic 20-kDa hGH possessed full growth-promoting activity in the weight gain test in hypophysectomized rats. However, contrary to previous work suggesting that native 20-kDa hGH lacks diabetogenic and insulin-like activities, biosynthetic 20-kDa hGH was found to have substantial diabetogenic activity when administered chronically to ob/ob mice and to possess approximately 20% the in vitro insulin-like activity of biosynthetic 22-kDa hGH on isolated epididymal adipose tissue of hypophysectomized rats. The diabetogenic and insulin-like activities of biosynthetic 20-kDa hGH cannot be ascribed to contamination of the hormone preparation with the 22-kDa form of hGH or with other diabetogenic or insulin-like pituitary peptides. Therefore, the results strongly suggest that diabetogenic and insulin-like activities are also intrinsic properties of the 20-kDa variant of hGH.A 20-kilodalton variant of human growth hormone (20-kDa hGH) has been isolated from human pituitary glands (1) and has been found to differ from the predominant 22-kilodalton form of hGH (22-kDa hGH) in that residues 32-46 are deleted (2, 3). The variant is produced in the human pituitary, because the nucleotide sequence encoding residues 32-46 is sometimes excised during the processing of hGH pre-mRNA to hGH mRNA (4). As a consequence, approximately 5-10%1 of the hGH present in the human pituitary is 20-kDa hGH (1). Physicochemical studies of 20-kDa hGH (3) suggest that its conformation is similar but not identical to that of 22-kDa hGH.The 20-kDa variant of hGH has been reported (1, 5) to have growth-promoting activity in the hypophysectomized rat equivalent to that of 22-kDa hGH. Moreover, its lactogenic activity in the pigeon crop sac assay equals that of 22-kDa hGH (1). On the other hand, Lewis et al. (5) reported that 20-kDa hGH lacked diabetogenic activity, in that it failed to produce hyperglycemia and glucose intolerance in dogs, when administered 10 hr prior to a glucose tolerance test. Also, this same group reported (6) that 20-kDa hGH lacked the acute insulin-like property of 22-kDa hGH, since the 20-kDa hGH variant failed to produce hypoglycemia and increase plasma free fatty acids when administered to hypophysectomized rats and did not increase glucose uptake or oxidation to CO2 by isolated rat adipose tissue wh...

Growth hormones. II. Structure—function relationships

“…They demonstrated that natural hGH and the Met-hGH derivative obtained by genetic engineering are similar in both weight gain and tibia tests in hypophysectomized rats. The same growth activity has been described by Kostyo et al (1985) for the 20,000-dalton Met-hGH.…”

“…hGH-N gene encodes for the sequence of the major form of hGH, the 22K, as well as the subsequent forms that are derived from it (see &dquo;Growth Hormone. I. Polymorphism&dquo;; Charrier and Martal, 1988) including the 20K variant from an alternative splicing of premessenger RNA (Wallis, 1980;De Noto et al, 1981), and a new natural 17.5K variant (Lecomte et al, 1987 (Frigeri et al, 1979;Kostyo et al, 1985). The second hGH-V gene seems to encode for a polypeptide of 191 amino acids experimentally transcribed in vitro, in which 13 amino acids differ from the 22K of hGH (Pavlakis et al, 1981).…”

“…Thus, although the 20K hGH is not totally deprived in vivo of insulin activity (Frigeri et al, 1979;Kostyo et al, 1985), it possesses a very small intrinsic insulin activity in vitro. Thus, peptide 32―46 is most likely involved in the insulin activity; and it should be noted that the basic amino acid at position 41 is particularly conservative in each GH (Table I) Tsushima and Friesen, 1973), rat (Robertson and Friesen, 1975), mouse (Talamantes, 1975;Colosi et aL, 1982) and hamster (Kelly et al, 1976;Southard et al, 1986) placental lactogen hormones.…”

“…Recently, Kawauchi and Yasuda (1987) Dayhoff (1976) (Martal, 1980 Kostyo et al, 1985). It also stimulates somatomedin production (Spencer et al, 1981 Lactogenic effects of the 20K hGH are similar to those observed in the presence of the 22K compound in the pigeon crop-sac assay (Lewis et al, 1978 In order to obtain further knowledge on the binding affinity of the 20 and 22K compounds for insulin-like receptors, Smal (1986) used hGH specific receptors isolated from cultured human lymphocytes (line IM-9) and rat adipocytes.…”

Biological Activities of Synthesized 20K and 22K hGH in Nb2 Bioassay and IM-9 Radioreceptor Assay.

“…But the insulin-like activity of 20K hGH has been reported to be absent or weak (Frigeri et al, 1979;Kostyo et al, 1985). The receptor reactivity of natural 20K hGH was reported to be about 30% of the standard hGH in IM-9 RRA (Hizuka et al, 1982).…”