Sucrase Is an Intramolecular Chaperone Located at the C-terminal End of the Sucrase-Isomaltase Enzyme Complex

Abstract

The sucrase-isomaltase enzyme complex (pro-SI) is a type II integral membrane glycoprotein of the intestinal brush border membrane. Its synthesis commences with the isomaltase (IM) subunit and ends with sucrase (SUC). Both domains reveal striking structural similarities, suggesting a pseudo-dimeric assembly of a correctly folded and an enzymatically active pro-SI. The impact of each domain on the folding and function of pro-SI has been analyzed by individual expression and coexpression of the individual subunits. SUC acquires correct folding, enzymatic activity and transport competence and is secreted into the external milieu independent of the presence of IM. By contrast, IM persists as a mannose-rich polypeptide that interacts with the endoplasmic reticulum resident molecular chaperone calnexin. This interaction is disrupted when SUC is coexpressed with IM, indicating that SUC competes with calnexin for binding of IM. The interaction between SUC and the membrane-anchored IM leads to maturation of IM and blocks the secretion of SUC into the external milieu. We conclude that SUC plays a role as an intramolecular chaperone in the context of the pro-SI protein.To our knowledge all intramolecular chaperones so far identified are located at the N-terminal end. SUC is therefore the first C-terminally located intramolecular chaperone in mammalian cells.Membrane and secretory proteins are subject to a complex array of cotranslational and post-translational processes that precede sorting to the organelle in which they exert their biological functions. The most crucial events take place in the ER 1 and include folding, subunit assembly, and in many cases oligomerization (for review see Ref. 1). These steps, individually or altogether, lead ultimately to the acquisition of the protein to a transport-competent conformation that enables its egress from the ER. A number of resident proteins of the ER, such as the molecular chaperones BiP and calnexin and protein disulfide isomerase, play primordial roles in these processes that vary from binding of unfolded proteins to the formation of disulfide bonds between individual subunits or within a single polypeptide chain (2-4). A large number of proteins assemble into homodimers, oligomers, or heterodimers preceding protein exit from the ER (5, 6). Correct protein folding of the individual monomers or subunits is required for optimal completion of these events. Many proteins are known not to go into dimers or heterodimers, such as the brush border proteins sucrase-isomaltase (SI) (7), angiotensin-converting enzyme (8), and maltase-glucoamylase (9). An interesting and common feature of these proteins is their composition of an even number of homologous domains. SI, for example, is an enzyme complex that is made of two subunits that share Ն 35% of amino acid sequence similarity and 41% of conserved sequences (10). Although no three-dimensional structure of this large protein is so far available, algorithmic predictions suggest that the two large domains are folded similarly and that...

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