Abstract: Methods and compositions are provided relating to production of recombinant protein in yeast. A modified PLAC4 is described where one or more mutations may be introduced into the Pribnow box-like sequences in the promoter. The modified promoter when placed upstream of a target gene in a vector causes a significant reduction of target gene expression in transformed bacteria but produces efficient expression of the target gene in yeast.

Abstract: Compositions and methods are provided for creating and identifying mutant carbohydrate-binding proteins that reversibly bind to carbohydrate substrates under conditions where the native protein remains bound. Examples of modified chitin-binding domains are provided which can be eluted from chitin in the presence of a reducing agent or at a pH within the range of 5-10.

Abstract: Compositions and methods are provided for reversibly binding chitin-binding domain (CBD) to a chitin or equivalent substrate under non-denaturing conditions. CBD from either prokaryotes or eukaryotes are modified for example, by random mutation, and screened to identify mutants that achieve this change in properties. Creating a modified CBD with an altered binding affinity for substrate provides new uses for CBD not previously possible with unmodified CBD that binds irreversibly to chitin.

Abstract: Methods and compositions are described that relate to obtaining concentrated preparations of secreted recombinant proteins. These proteins are expressed in the form of fusion proteins with a chitin-binding domain (CBD). The fusion proteins are capable of being concentrated in the presence of chitin. Also described is: a shuttle vector that includes a modified LAC4 promoter; a chitinase-negative host cell; a CBD capable of eluting from chitin under non-denaturing conditions; and sterilized chitin, which can be optionally magnetized for facilitating recovery of recombinant protein.

Abstract: Compositions and methods are provided for reversibly binding chitin binding domain (CBD) to a chitin or equivalent substrate under non denaturing conditions. CBD is modified preferably by a mutation to achieve this change in properties. In one embodiment, an aromatic amino acid residue located in the binding cleft of the CBD was altered resulting in reversible binding affinity for substrate in select conditions. Creating a modified CBD with an altered binding affinity for substrate provides new uses for CBD not previously possible with unmodified CBD which binds irreversibly to chitin.

Abstract: Methods and compositions are provided relating to production of recombinant protein in yeast. A modified PLAC4 is described where one or more mutations may be introduced into the Pribnow box-like sequences in the promoter. The modified promoter when placed upstream of a target gene in a vector causes a significant reduction of target gene expression in transformed bacteria but produces efficient expression of the target gene in yeast.