HNL from Sorghum bicolor

SfeHNL (Sordan 79 variety) is a heterotetrameric enzyme (a/P)2. Both a- (33 kDa) and P-subunits (23 kDa) are glycosylated and linked by two disulfide bridges [31, 33]. The a- and p-subunit are synthesized in one pre-protein, which is cleaved proteolytically after protein biosynthesis (Fig. 2.2.3.3). 

Carboxypeptidase side-activity Since the protein sequence and 3D structure of 

Meanwhile the 3D structure of SbHNL was solved [27], showing the expected high similarity to the structure of GPU from wheat [41]. Both enzymes use the same active-site residues. However, in SfeHNL a unique two-amino acid deletion 

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A suspension of Avicel cellulose (0.5 g) in 0.05 mmol/L phosphate buffer (pH 4.5, 10 mL) containing ammonium sulfate (4.72 g) was stirred for 1 h, and a solution of (S)-HNL from Sorghum bicolor (50 pL, 1000 IU/mL, specific activity 70 IU/mg) was added. The mixture was stirred at room temperature for 10 min and filtered, and the immobilized enzyme was suspended in diisopropyl ether (10 mL). After addition of aldehyde (2 mmol) and dry liquid HCN (300 pL, 7.5 mmol), the mixture was stirred until all aldehyde had reacted. After removal of the immobilized enzyme, the filtrate was concentrated to yield the crude cyanohydrin. 

Another option for the identification of the gene is the immunoscreening of cDNA libraries with antibodies produced specifically using the respective purified proteins (e.g., PP HNL from Hevea brasiliensis [80], Sl HNL from Sorghum bicolor [61]). 

The HNL from Sorghum bicolor was shown to consist of one major and two minor isoenzymes [57]. Crystallographic studies on the three isoenzymes were reported and preliminary x-ray investigations of crystallized forms were published [65]. 

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