Subtilisin inhibitor

A large number of potential reversible protease inhibitors exist (Laskowski Kato, 1980). Protein protease inhibitors like Strepromyces Subtilisin Inhibitor (SSI) (Hiromi et al, 1985) and Chymotrypsin Inhibitor (CI-2) (Jonassen, 1980 and McPhalen James, 1988) are known to be very strong inhibitors with inhibition constants at or below 10"10 M. 

Hiromi, K Akasaka, K Mitsui, Y Tonomura, B Murao, S (eds) (1985) Protein Protease Inhibitor - The Case of Streptomyces Subtilisin Inhibitor. Elsevier Amsterdam - Oxford - New York. 

Aspartate transcarbamylase regulatory domains 1 and 2 Streptomyces subtilisin inhibitor Glutathione reductase domain 3 Thermolysin domain 1... 

Hexokinase Pyruvate kinase Adenylate kinase Phosphoglycerate kinase Phosphofructokinase Protease inhibitors Pancreatic trypsin inhibitor Soybean trypsin inhibitor Streptomyces subtilisin inhibitor Nucleases... 

Contacts with the catalytic residues, in combination with hydrophobic interactions, are also observed in the complex of an insect a-amylase with the Ragi bifunctional a-amylase/trypsin inhibitor (RBI) [174]. Conversely, the mechanism of inhibition of barley a-amylase by the barley a-amylase/subtilisin inhibitor (BASI) did not involve direct contact between inhibitor residues and the catalytic site [175]. The inhibitor sterically blocks the catalytic site, but does not extend into it. A cavity is created, which is occupied by a calcium ion coordinated by water-mediated interactions with the catalytic residues. 

Data are adapted from compilations of Dure et at. (1989). O, high osmoticum (PEG or salt) D, desiccation C, cold W, wounding H, heat , untested or unknown. Sd, seed St, stem Pase, protease ASI, a-amylase/subtilisin inhibitor WGA, wheat germ agglutinin RNP, ribonuclear protein not organ specific. Table updated from Skriver Mundy (1990). 

Fig. 3. Translation products of barley aleurone layer mRNAs. Lanes 1-4, total products from layers treated in water (control), 1 jxm ABA, 1 pM GA3, or both hormones, respectively. The rab 6 protein is marked with an arrow. Lanes 5-8, immunoprecipitation of a-amylase (upper bands) and the a-amylase/subtilisin inhibitor (lower bands) from the translation products shown in lanes 1-4. mRNA extraction, translation and immunoprecipitation were performed as previously described (Mundy etal., 1986).

Leah, R. Mundy, J. (1989). The bifunctional a-amylase/subtilisin inhibitor of barley Nucleotide sequence and patterns of seed-specific expression. Plant Molecular Biology 12, 673-82. 

The purification and characterization of BmSI-7 and BmSI-6, two subtilisin inhibitors from Boophilus microplus (BmSI) was reported by Sasaki et al. (2008). The inhibitors were found to exhibit significant inhibition on the activity of purified Prl proteases from M. anisopliae. 

Sasaki, S. D., de Lima, C. A., Lovato, D. V., Juliano, M. A., Torquato, R. J. S., and Tanaka, A. S. (2008). BmSI-7, a novel subtilisin inhibitor from Boophilus microplus, with activity toward Prl proteases from the fungus Metarhizium anisopliae. Experimental Parasitology, 118, 214-220. 

Nuclear Magnetic Resonance Studies on Streptomyces Subtilisin Inhibitor and its Complexes with Proteinases... 

Fig. 2.1 Structure of SSI subunit. (Reproduced with permission from Y., Mitsui, et al Protein Protease Inhibitor-The Case of Streptomyces Subtilisin Inhibitor (SSI) (K. Hiromi et al. ed.), p. 174, Elsevier, Amsterdam (1985)).

Hordeum vulgare (barley) (Poaceae)[seed] BASI (barley a-Amylase Subtilisin inhibitor) (177 aa 20 kDa ) Subtilisin [0.2 nM] (V67-A68) (also inhibits a-Amylase) [372- 375]... 

Oryza sativa (rice) (Poaceae)[seed] Rice a-Amylase-Subtilisin Inhibitor (RAS1) (176 aa 19 kDa 4 Cys 2 S-S) (minor variants also) Subtilisin (A67-A68) ( also inhibits a-Amylase) [389]... 

Triticum aestivum (wheat) (Poaceae) [seed] WASI (Wheat a-Amylase Subtilisin inhibitor) (180 aa 20 kDa Kunitz-related protein 4 Cys 2 S-S) Subtilisin ( a-Amylase) [434, 435]... 

Kainosho M, Tsuji T, Assignment of the three methionyl carbonyl carbon resonances in Streptomyces subtilisin inhibitor by a carbon-13 and nitrogen-15 double-labeling technique. A new strategy for structural studies of proteins in solution, Biochemistry, 21 6273-6279, 1982. 

Another important area of dynamic studies in biological samples is the effect of hydration upon molecular mobility in proteins and carbohydrates. The reason for these studies is primarily that protein dynamics, in particular, are crucial to their function, and so examining factors, such as the degree of hydration, that affect their dynamics is very important. However, it is obviously near-impossible to study dynamics in aqueous solution as a function of degree of hydration, and, since most proteins are not soluble in nonaqueous solvents, solid-state studies must be used. The motions at three methionine (Met) residues in Streptomyces subtilisin inhibitor (SSI) were studied with 2H NMR using a sample in which the Met residues at two crucial enzyme recognition sites (PI and P4) were specifically deuterated, along with one in the hydrophobic core.114 The motions of the Met side-chains were then examined... 

Another major class of proteins, the a has approximately equal amounts of a-helical and 0 sheet structure. Often there is a cluster of helices at one or both ends of a single 0 sheet. 0 structure is generalh found as antiparallel strands. One particular type oi a - - 0 structure is termed the open-face sandwich 0 structure. The subtilisin inhibitor has a five-stranded 0 structure with two a helices protecting its open face. 

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