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Rapid Maturation of Glycoprotein Hormone Free -Subunit (GPH) and GPH Homodimers

Heidelberg University Biochemistry Center (BZH) (J.-M.K., J.R., V.S., W.E.M.), 69120 Heidelberg, Federal Republic of Germany; Institute for Biomedical Aging Research, Austrian Academy of Sciences (P.B.), Innsbruck A6020, Austria; and Hormone Biochemistry Laboratory, Institute of Self Organizing Systems and Biophysics (V.S.), North-Eastern Hill University, Permanent Campus, Shillong 793022, Meghalaya, India

Address all correspondence and requests for reprints to: Wolfgang E. Merz, Ph.D., Heidelberg University Biochemistry Center (BZH), Im Neuenheimer Feld 328, 69120 Heidelberg, Federal Republic of Germany. E-mail: wolfgang.merz{at}bzh.uni-heidelberg.de.

The dynamics of glycoprotein hormone -subunit (GPH) maturation and GPH homodimer formation were studied in presence (JEG-3 choriocarcinoma cells) and absence (HeLa cells) of hCGß. In both cases, the major initially occurring GPH variant in [³⁵S]Met/Cys-labeled cells carried two N-glycans (M_{r app} = 22 kDa). Moreover, a mono-N-glycosylated in vivo association-incompetent GPH variant (M_{r app} = 18 kDa) was observed. In JEG-3 cells the early 22-kDa GPH either associated with hCGß, or showed self-association to yield GPH homodimers, or was later converted into heavily glycosylated large free GPH (M_{r app} = 24 kDa). Micro-preparative isolation of intracellular GPH homodimers of JEG-3 cells and their conversion by reduction revealed that they consisted of 22-kDa GPH monomers and not of large free GPH. In HeLa cells, the large free GPH variant was not observed, whereas GPH homodimers were present. Intracellularly, early GPH homodimers (35 kDa) and late variants (JEG-3: 44 kDa, HeLa: 39 kDa) were found. Both cell types secreted 45 kDa GPH homodimers. Large free GPH and GPH homodimers were more rapidly sialylated than hCG ß-heterodimers indicating a sequestration mechanism in the secretory pathway. In GPH homo- as well as hCG ß-heterodimers the subunit interaction site, located on loop 2 of GPH (amino acids 33–42), became immunologically inaccessible indicating similar spatial orientation of GPH in both types of dimers. The studies demonstrate the formation, in vivo dynamics of GPH homodimers, and the pathways of the cellular metabolism of variants of GPH, monoglycosylated GPH and large free GPH.

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