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An active site is the region of an enzyme where a substrate binds and undergoes a chemical reaction. In a protein enzyme, it includes the amino acid residues that contact the substrate and those that directly assist catalysis. These roles can overlap: an amino acid side chain can both orient a substrate and participate in chemistry. The active site occupies only part of the folded protein; the remaining amino acid residues help establish its protein structure, stability and movement. An active site need not be a consecutive stretch of protein primary structure: protein folding can bring distant amino acid residues together. A ribozyme instead supplies an RNA active site. See Alberts et al., Molecular Biology of the Cell, Protein Function: binding sites and catalytic sites.

Binding and catalysis answer different questions. A protein binding site recognizes shape and chemistry through such interactions as hydrogen bonds, electrostatics and the hydrophobic effect. At an active site, appropriate positioning also helps a substrate reach the transition state of a chemical reaction. Some amino acid side chains transfer a proton; others participate transiently in a covalent bond. The enzyme-substrate complex is the associated enzyme and substrate, before conversion to a reaction product. Merely accumulating this complex does not establish rapid catalysis: stabilizing the bound substrate without sufficiently stabilizing the transition state can leave a large barrier. The important complementarity is chemical as well as geometric. See Geoffrey M. Cooper, The Cell, The Central Role of Enzymes as Biological Catalysts: substrate binding and transition-state stabilization.

The cartoon makes a useful experimental distinction. Changing a contacting amino acid residue might weaken substrate binding, change the reaction rate, or do both. Changing a distant amino acid residue can also disrupt the active site by changing protein folding. Therefore a loss of activity after a mutation alone does not locate a catalytic group: compare binding, protein structure and enzyme kinetics as well. This is an interpretation guide, not a report of a particular mutation experiment.

A concrete example is hexokinase: it transfers the terminal phosphate group of ATP to the $6$-position hydroxyl group of a sugar such as glucose. Specificity need not mean only one acceptable substrate: the hexokinase reaction class includes several possible acceptor sugars. Which are used, and at what reaction rates, depends on the particular enzyme and conditions. See Swiss Institute of Bioinformatics, ENZYME entry EC 2.7.1.1: hexokinase reaction and acceptor range.

Bennett and Steitz compared yeast hexokinase A crystallized with glucose against hexokinase B crystallized without glucose. X-rays scattered by the crystals supplied diffraction measurements from which the authors reconstructed protein structure; comparing coordinates located movement, rather than measuring a reaction rate. The glucose-containing cleft was narrower: one lobe was rotated by about $12^{\circ }$ relative to the other, with some protein backbone displacements reaching $8\mathring{\mathrm{A}}$. This makes the active site a moving arrangement of interacting groups, rather than a permanent hole. See William S. Bennett Jr. and Thomas A. Steitz, Glucose-induced conformational change in yeast hexokinase, PNAS 75 (1978), 4848–4852.

The structural comparison has two explicit limitations: different forms of hexokinase and different crystal environments were compared, and static coordinates do not record the order of binding and movement. Closure is consistent with induced fit, but those structures alone cannot exclude conformational selection. The solution experiment under induced fit addresses whether compaction also occurs outside a crystal.

A beta sheet consists of extended stretches of protein backbone lying alongside one another, joined by backbone hydrogen bonds. The stretches can belong to different portions of the same polypeptide chain or to different chains.

Each extended stretch is a beta strand. Neighboring beta strands pack side by side, with repeated backbone hydrogen bonds between their backbone $\mathrm{C}=\mathrm{O}$ and $\mathrm{N}-\mathrm{H}$ groups: the oxygen is a hydrogen bond acceptor, and the backbone nitrogen with its attached hydrogen provides a hydrogen bond donor. These are interactions of the shared protein backbone, not merely contacts between amino acid side chains. Unlike an alpha helix, a beta sheet joins adjacent extended stretches rather than coiling one stretch around an axis. See Clark, Douglas and Choi, Biology 2e, 3.4: backbone groups in secondary structure.

Read each beta strand in the sequence direction from the N-terminus toward the C-terminus, even when it is an internal part of a longer polypeptide chain. In a parallel beta sheet, neighboring beta strands run in the same N→C direction; in an antiparallel beta sheet, they run in opposite N→C directions. A three-strand example can therefore have arrows right–right–right or right–left–right. A beta sheet can also mix the two arrangements. Direction does not identify chain membership: a single polypeptide chain can fold back to supply several beta strands, and different polypeptide chains can contribute to one beta sheet. See Alberts et al., Molecular Biology of the Cell: beta-sheet strand arrangements.

The pleated appearance comes from the geometry of the protein backbone. Successive amino acid residues carry amino acid side chains on alternating faces of an ideal beta strand, conventionally drawn above and below the sheet. The attachment positions are the amino acid alpha carbons. Thus a six-position cartoon has groups 1, 3 and 5 on one face and 2, 4 and 6 on the other. This alternation is a spatial pattern, not a rule that alternating positions must contain identical amino acids. See Clark, Douglas and Choi: side chains above and below the pleats.

Real beta sheets commonly twist and curve; pleating does not make them perfectly flat plates. Which beta strands are neighbors, how connecting regions join them, and how their faces pack are features of the three-dimensional protein structure. An alternating pattern in the protein primary structure alone neither establishes a beta sheet nor specifies a unique fold independently of surroundings and interacting molecules. See Breda et al., Protein Structure, Modelling and Applications: extended strands, sheet twist and three-dimensional organization and protein folding.

Why this arrangement is useful: repeated backbone hydrogen bonds organize extended chain regions into a framework that can contribute to the interior of a folded protein. Amino acid side chains and connecting loops give that framework chemical variety and can shape a protein binding site for a ligand. Its attractive feature is the combination of a reusable backbone pattern with variable exposed groups: similar beta sheets can support different recognition surfaces. See Alberts et al.: beta-sheet cores and loops that form binding sites.

Historically, Linus Pauling and Robert B. Corey published The Pleated Sheet, a New Layer Configuration of Polypeptide Chains in PNAS 37, 251–256 (1951). They reasoned from peptide bond geometry and possible backbone hydrogen bonds, proposing an idealized layer rather than measuring every modern beta sheet. Their initial model should not be identified with all present-day beta sheet geometries; they published further pleated-sheet configurations later in 1951. See Pauling and Corey: original 1951 paper and scan, ^[ref], and Pauling and Corey, Configurations of Polypeptide Chains With Favored Orientations Around Single Bonds: Two New Pleated Sheets, PNAS 37, 729–740 (1951).

A protein binding site is a region of a protein structure that interacts with a ligand. Its arrangement of amino acid residues helps determine recognition; connecting loops supported by a beta sheet can contribute to it. An enzyme's active site binds a substrate and accelerates a chemical reaction. See Alberts et al.: protein binding sites and active sites.

The protein primary structure specifies the ordered amino acid residues of each polypeptide chain, joined through peptide bonds. A chain is conventionally written from its N-terminus to its C-terminus. In the broader chemical sense, a complete protein primary structure also specifies covalent bond crosslinks between chain positions, including protein disulfide bonds; a bare sequence string leaves that information unstated. This describes chemical connectivity rather than the spatial arrangement of protein tertiary structure or protein quaternary structure. See IUPAC Gold Book: primary structure, sequence and chain crosslinking and IUPAC-IUB, 3AA-16: writing residue sequences from the amino end. Some conventions reserve protein primary structure for sequence alone and call the complete chemical connectivity the covalent structure; the distinction should be stated when comparing descriptions. See IUPAC-IUB polypeptide-conformation recommendations, section 6.1: the narrower sequence-only convention.

In this toy example M denotes methionine, A alanine and G glycine. Both polypeptide chains have six amino acid residues, five peptide bonds, an M at the N-terminus and a G at the C-terminus. Their amino acid composition is identical, yet position 2 is A in MAGAMG and M in MMAAGG. There are $6!/(2!2!2!)=90$ distinct orders with these counts, a multinomial coefficient. Even fixing those two end residues leaves $4!/(1!2!1!)=12$ orders. The counts constrain the answer without determining it.

For multiple polypeptide chains, record each sequence separately and specify any protein disulfide bonds between cysteine positions. Mature insulin, for example, has an A chain of 21 amino acid residues and a B chain of 30 amino acid residues. Their combined 51 residues give $(21-1)+(30-1)=49$ peptide bonds, rather than the 50 of one unbroken 51-residue chain. See Cooper, The Cell: insulin's two chains and cysteine crosslinks, figure 2.16. A protein disulfide bond joins side groups; it does not concatenate the C-terminus of one chain with the N-terminus of another. The three insulin protein disulfide bonds connect A6 to A11 within the A chain, and A7 to B7 and A20 to B19 between chains. Positions are numbered from each chain's N-terminus. See Replacement of the interchain disulfide bridge-forming amino acids A7 and B7 by glutamate impairs the structure and activity of insulin: native disulfide positions.

Why keep this information? Protein primary structure lets us distinguish changes of chemical identity from changes of shape, and compare corresponding positions between proteins. Protein folding can bring distant amino acid residues together at a protein binding site while preserving their sequence. The attractive idea is that a readable linear record can constrain a complicated three-dimensional object; it is not, by itself, a guarantee of a unique fold in every environment. See protein folding and the ribonuclease A refolding experiment. Merely bending an intact chain preserves its protein primary structure; hydrolysis of a peptide bond, or changing which positions a protein disulfide bond connects, changes its chemical connectivity.