Only a mutated CH1 domain lacking Cys128 serves as a proper template for two Cys214 to form the alternative interchain disulfide that generates the LC2homo-dimer. An important feature of the unique tetrameric IgG structure described here is its ability to be secreted. can generate more than one functional structure that is considered native by the cellular quality control system. Keywords:IgG, assembly, disulfide bonds, secretion == 1. Introduction == Disulfide bonds within proteins are responsible for stabilizing folded structure and disulfide bonds between polypeptides are responsible for covalent assembly. Immunoglobulins (Igs) serve as an excellent model to study disulfide bond formation because, based on a large database of Ig sequences Gastrofensin AN 5 free base and structures, it is obvious that these oligomeric proteins contain highly conserved disulfide bonds within their heavy chains (HC) and light chains (LC) and between HC and LC. A single disulfide bond within each variable or constant domain name of HC or LC stabilizes the typical sandwich fold of these domains. In addition, two types of interchain disulfide bonds stabilize the tetrameric structure of functional Igs; one that links two HC to each other and another that bridges each HC to LC (Edelman and Gall 1969). Assembly of LCs and HCs is initiated by non-covalent interactions between their respective VLand VHvariable Gastrofensin AN 5 free base regions (Percy et al. 1975;Secher et al. 1977;Hamel et al. 1984;Chothia et al. 1985;Hamel et al. 1987). This is followed by hydrophobic conversation between CLand CH1, the constant domains of LC and HC, respectively (Janeway 1994), which is usually eventually stabilized by a covalent interchain disulfide between these two constant domains (Hamel et al. 1987;Hendershot et al. 1987;Leitzgen et al. 1997). The CH1- CLinterchain disulfide forms between Cys214 of CL(or Cys215 of CL) and the most N-terminal among three cysteines in CH1. In mouse IgG2b, the interchain disulfide with LC is usually created TMPRSS2 by Cys128, which is located in the loop connecting the first and second -strands of CH1, as suggested by both conserved cysteine positions (Amzel and Poljak 1979) and the functions assigned to them in the CH1 of the mouse 2b HC (hereafter referred to as ) (Rose et al. 1993). The conversation of this loop Gastrofensin AN 5 free base with CLappears to play a role in the assembly of HC with LC. Known exceptions are IgG1 and human IgA2(n), in which the LC-interacting cysteine is the most C-terminal, hinge domain-proximal cysteine in CH1 or the hinge-proximal cysteine in the CH2 domain name (Chintalacharuvu et al. 2002), respectively. The usual assembly pathway of IgG2b in Ig generating cells is Gastrofensin AN 5 free base usually HC-LC HC2LC HC2LC2(Scharff et al. 1970). An alternative IgG2b assembly pathway, which predominates in COS-7 cells, is usually HC HC2 HC2LC HC2LC2(Elkabetz et al. 2005). This pathway is also followed during assembly of other Ig isotypes (Scharff et al. 1970). These alternate pathways show that formation of the interchain disulfide bond is not an autonomous feature of the subunits, but is usually influenced by the cellular environment, a conclusion that was also reached recently by (Chintalacharuvu et al. 2007). Regardless of the assembly pattern, most naturally occurring Igs are secreted either as HC2LC2covalent tetramers or as HC2LC accompanied by one non-covalent LC, but not as HC2accompanied non-covalently by two LCs. This phenomenon may reflect quick covalent assembly of HC2into HC2LC and unfavorable cooperativity in the covalent assembly of the second LC with the HC2LC intermediate (Kazin and Beychok 1978). It is also plausible that a disulfide bond between HC2and at least one of the two accompanying LCs is required to pass the scrutiny of the quality control mechanisms operating in the endoplasmic reticulum (ER). However, while the covalent assembly via disulfides is the norm, the presence of exceptions like human IgA2m(1), which is usually secreted largely without a covalent bond between HC and LC (Chintalacharuvu et al. 2002), shows that alternative structures are possible. In addition to the vital contribution of.